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Biomedical subjects

M Zhang

Publications and source records attributed to M Zhang.

At least 559 records · Page 31Linked to original sources
PubMed1999

[Coprecipitation for preconcentration and determination of trace cadmium in solid waste phosphogypsum by FAAS].

CuS was used as a collector precipitate to preconcentrate and separate trace cadmium in phosphogypsum. The optimum coprecipitation conditions were studied, it was found that Cd could be quantitatively coprecipitated with CuS in sample solution at pH 5.8-7.0 and that 100 microg of Cu was enough to coprecipitate Cd in 50 mL of sample solution. No interferences of coexistent elements were found. A sample was detected for 6 times. The RSD was 7.97% and the recovery was between 88% and 110%. The method was rapid, simple and satisfactory for detecting Cd in phosphogypsum.

Cadmium↗
PubMed1999

[Study on the synergistic coloration of tin (II) with pyrocatechol violet and rhodamine B and its application].

In the presence of PVA, a sensitive color reaction of tin (I ) on pyrocatechol violet and rhodamine B has been studied. In a NaAc-HAc (pH = 5.0) medium, tin(II) ions form an ions-associate complex with pyrocatechol violet and rhodamine B. The composition of the complex is Sn (II): PV : RhB = 1 : 2: 4. The maximum absorption of the ions-associate complex is at 560 nm with an apparent molar absorptivity of 1.18 x 10(5) L x mol(-1) x cm(-1). Beer's law is obeyed in the range of 0-15 microg/25 mL of tin. The method is highly selective and has been applied to the determination of trace tin in some objects with satisfactory results.

Benzenesulfonates↗
PubMed1998

A peptide corresponding to residues Asp177 to Asn208 of human cyclin A forms an alpha-helix.

Cyclins are essential activators of eukaryotic cell cycle-regulating enzymes called cyclin-dependent kinases (CDKs). The binding of cyclins to CDKs is mediated by a structural motif comprising a five-helix bundle called the cyclin fold and an additional helix (the N-terminal alpha-helix) located N-terminal to the cyclin fold. In this work, we examine, using CD and NMR spectroscopy, the structure of a 32-residue synthetic peptide derived from the segment (Asp177 to Asn208) corresponding to the N-terminal alpha-helix of human cyclin A. CD spectroscopic analysis of the peptide revealed that trifluoroethanol (TFE) can induce the peptide to assume a stable alpha-helix conformation. Two-dimensional 1H NMR spectroscopy showed that the alpha-helix is formed by the Asp181 to Cys193 segment of the peptide. The alpha-helical structure of the peptide in the TFE/H2O cosolvent was found to be identical to that in the crystal structure of intact cyclin A. Taken together, these results suggest that the N-terminal alpha-helix of cyclins may exist as an independent structural unit that plays essential functional roles in activating CDKs.

Amino Acid Sequence↗
PubMed1998

Protein inhibitor of neuronal nitric-oxide synthase, PIN, binds to a 17-amino acid residue fragment of the enzyme.

Neuronal nitric-oxide synthase (nNOS) is the primary nitric oxide (NO) regulator in neurons. The activity of the enzyme is inhibited by a protein inhibitor called PIN. We were able to purify large quantities of PIN overexpressed in bacterial cells. Analytical ultracentrifugation and chemical cross-linking studies showed that PIN exists as a monomer at low concentrations. The protein forms a high order aggregate at elevated concentrations. We have shown, using NMR spectroscopy, that the previously identified PIN-binding domain (PINB) of nNOS (residues 161-245) adopts a random coil structure in solution. By titrating 15N-labeled PINB with unlabeled PIN, the PIN-binding region of nNOS was precisely mapped to a 17-residue peptide fragment from Met-228 to His-244 of nNOS. NMR titration experiments also showed that PIN binds to nNOS with a 1:2 stoichiometry. A synthetic peptide corresponding to the identified PIN-binding region of nNOS was used to study the interaction between PIN and nNOS in detail. The functional implications of the results obtained from this study are discussed.

Amino Acid Sequence↗
PubMed1998

Synthesis and biological properties of new 2beta-alkyl- and 2beta-aryl-3-(substituted phenyl)tropane derivatives: stereochemical effect of C-3 on affinity and selectivity for neuronal dopamine and serotonin transporters.

In our efforts to identify molecules that might act as cocaine antagonists or cocaine partial agonists, we have been involved in efforts to further elucidate the nature of cocaine's binding to the dopamine transporter (DAT) through strategic modifications of its structure. In the case of the substituent located at the 2-position of the tropane ring, studies have revealed the ability of the transporter to accommodate groups of diverse structure, including ester, ketone, alkyl, alkenyl, heterocyclic, and aryl substituents, without loss of DAT binding affinity. In the present study, we report our results pertaining to the ability of the DAT to accommodate the WIN-type structures possessing alkyl or aryl groups at the 2-position and which adopt either a chair or a boat conformation of the tropane ring. Moreover, we discuss the influence of the stereochemistry of these compounds in their selectivity for the DAT versus the serotonin transporter (5HTT). Additionally, we point out the importance of using Ki values rather than IC50 values when making such comparisons of transporter selectivity. One of the most interesting compounds identified in the present work is a 2, 3-diaryltropane 22 in a boat conformation that is highly selective (69-fold) for the DAT over the 5HTT. The ability to prepare this compound as well as related structures by our oxidopyridinium betaine-based dipolar cycloaddition strategy further underscores the versatility of this particular chemical approach to the preparation of diverse tropane analogues. The use of the optically pure olefin p-tolyl vinyl sulfoxide as the dipolarophile in this reaction allows access to these novel tropanes in nonracemic form.

Animals↗
PubMed1998

The longer isoform and Cys-1 disulfide bridge of rat surfactant protein A are not essential for phospholipid and type II cell interactions.

Rat SP-A is a heterooligomer of two closely related isoforms, that requires interchain disulfide linkage for several functions including SP-A-mediated phospholipid vesicle aggregation and modulation of surfactant secretion and uptake by isolated alveolar type II cells. While the Cys6 disulfide bond of rat SP-A is known to be critical for function, the importance of the second interchain disulfide linkage within the N-terminal Isoleucine-3-Lysine-Cysteine-1 (IKC) sequence of the alternatively processed isoform is not clear. To examine the role of the Cys-1-dependent multimerization in SP-A function, we disrupted the Cys-1 disulfide bond by deletion of the IKC sequence (SP-Ahyp, DeltaIKC) or the substitution Cys-1 to Ser (SP-Ahyp,C-1S) in mutant recombinant proteins produced in insect cells. N-terminal sequence analyses revealed that the mutations influenced signal peptidase cleavage specificity, resulting in an increase in the abundance of the longer isoform of SP-Ahyp,C-1S and in N-terminal truncation of a fraction of the SP-Ahyp,DeltaIKC polypeptides at Gly8. On nonreducing SDS-PAGE analysis, both mutant proteins migrated as monomers and dimers but not the higher multimers that are characteristic of the wild-type recombinant protein (SP-Ahyp). Cross-linking analyses demonstrated that the association between trimeric SP-A subunits remained intact despite disruption of the Cys-1 bond. The SP-Ahyp,C-1S eluted in the same volume as SP-Ahyp from the gel sizing column with an apparent mass of 440 kDa, indicative of association of at least 9-10 monomers. The phospholipid binding and aggregation activities of the SP-Ahyp,C-1S were approximately 75% and 55% of the SP-Ahyp, respectively, but the SP-Ahyp,DeltaIKC was functionally comparable to SP-Ahyp. Similarly, both mutant proteins regulated the secretion and uptake of surfactant from isolated type II cells, but the SP-Ahyp,DeltaIKC was somewhat more potent than the SP-Ahyp,C-1S. Competitive binding to the SP-A receptor on type II cells was reduced by both Cys-1 mutations. We conclude that neither Cys-1-dependent multimerization nor the longer SP-A isoform is absolutely required for oligomeric association of trimeric SP-A subunits, SP-A/phospholipid interactions, or the regulation of surfactant secretion or uptake from type II cells by rat SP-A.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1998

Fetuin (alpha2-HS-glycoprotein) opsonizes cationic macrophagedeactivating molecules.

Macrophages become activated by bacterial endotoxin (lipopolysaccharide) and other stimuli to release proinflammatory cytokines and NO. To prevent release of toxic or potentially lethal quantities of these factors, the state of macrophage activation is counter-regulated by anti-inflammatory mediators (e.g., glucocorticoid hormones, interleukin 10, and transforming growth factor type beta). Fetuin, a negative acute-phase protein, recently was implicated as an anti-inflammatory mediator, because it is required for macrophage deactivation by spermine. In the present studies, we found that fetuin is necessary for macrophages to respond to CNI-1493, a tetravalent guanylhydrazone inhibitor of p38 mitogen-activated protein kinase phosphorylation. Fetuin dose-dependently increases macrophage uptake of CNI-1493, which can be specifically inhibited by anti-human fetuin antibodies. Anti-human fetuin antibodies render primary human peripheral blood mononuclear cells insensitive to deactivation by CNI-1493. Thus, macrophages use fetuin as an opsonin for cationic-deactivating molecules, both endogenous (e.g., spermine) and pharmacologic (e.g., CNI-1493). This role of fetuin as an opsonic participant in macrophage-deactivating mechanisms has implications for understanding and manipulating the innate immune response.

Amino Acid Sequence↗
PubMed1998

Randomised placebo-controlled trial of use of topical recombinant bovine basic fibroblast growth factor for second-degree burns.

BACKGROUND: Wound healing is a dynamic process that could be accelerated by growth factors. We investigated the effect of recombinant bovine basic fibroblast growth factor (rbFGF) on burn healing in a randomised placebo-controlled trial. METHODS: We recruited 600 patients with superficial or deep second-degree burns. Patients received 150 AU/cm2 daily topical rbFGF (n=300) or placebo (n=300) plus vehicle. We assessed healing by photography, punch-biopsy, and clinical examination. FINDINGS: All patients treated with rbFGF had faster granulation tissue formation and epidermal regeneration than those in the placebo group. Superficial and deep second-degree burns treated with rbFGF healed in a mean of 9.9 (SD 2.5) days and 17.0 (4.6) days, respectively, compared with 12.4 (2.7) and 21.2 (4.9) days (p=0.0008 and p=0.0003, respectively). No adverse effects were seen locally or systemically with rbFGF. INTERPRETATION: rbFGF effectively decreased healing time and improved healing quality. Clinical benefits would be shorter hospital stays and the patient's skin quickly becoming available for harvesting and grafting.

Administration, Cutaneous↗
PubMed1998

Overlapping functions of E- and P-selectin in neutrophil recruitment during acute inflammation.

Selectin adhesion molecules mediate leukocyte rolling on activated endothelium, a prerequisite to leukocyte accumulation at sites of inflammation. The precise role of each selectin (E-, P-, and L-) in this process is unclear and may vary depending on the particular inflammatory stimulus, vascular bed, leukocyte subset, and species; most data suggest discrete functional roles for each selectin. To define the relative roles of E- and P-selectin in mediating neutrophil accumulation in acute dermal inflammation, mice genetically deficient in E-selectin, P-selectin, or both E- and P-selectin were injected intradermally with zymosan. Luminal endothelial expression of E- and P-selectin in response to zymosan was documented in wild-type mice by intravenous administration of fluorochrome-labeled anti-E- and anti-P-selectin antibodies. In mice deficient in E- or P-selectin, neutrophil accumulation was unchanged or only subtly reduced relative to wild-type control mice. In mice deficient in both E- and P-selectin, neutrophil accumulation was significantly reduced (87% at 4 hours and 79% at 8 hours). These data demonstrate that, in this model of acute inflammation, there is considerable overlap in the functions of E- and P-selectin; loss of both selectins was required to impair neutrophil accumulation.

Animals↗
PubMed1998

Toll-like receptor-2 mediates lipopolysaccharide-induced cellular signalling.

Vertebrates and invertebrates initiate a series of defence mechanisms following infection by Gram-negative bacteria by sensing the presence of lipopolysaccharide (LPS), a major component of the cell wall of the invading pathogen. In humans, monocytes and macrophages respond to LPS by inducing the expression of cytokines, cell-adhesion proteins, and enzymes involved in the production of small proinflammatory mediators. Under pathophysiological conditions, LPS exposure can lead to an often fatal syndrome known as septic shock. Sensitive responses of myeloid cells to LPS require a plasma protein called LPS-binding protein and the glycosylphosphatidylinositol-anchored membrane protein CD14. However, the mechanism by which the LPS signal is transduced across the plasma membrane remains unknown. Here we show that Toll-like receptor 2 (TLR2) is a signalling receptor that is activated by LPS in a response that depends on LPS-binding protein and is enhanced by CD14. A region in the intracellular domain of TLR2 with homology to a portion of the interleukin (IL)-1 receptor that is implicated in the activation of the IL-1-receptor-associated kinase is required for this response. Our results indicate that TLR2 is a direct mediator of signalling by LPS.

Binding Sites↗
PubMed1998

Characterization of siglec-5, a novel glycoprotein expressed on myeloid cells related to CD33.

We describe the characterization of siglec-5 (sialic acid-binding Ig-like lectin-5), a novel transmembrane member of the immunoglobulin superfamily, highly related to the myeloid antigen, CD33. A full-length cDNA encoding siglec-5 was isolated from a human activated monocyte cDNA library. Sequencing predicted that siglec-5 contains four extracellular immunoglobulin-like domains, the N-terminal two of which are 57% identical to the corresponding region of CD33. The cytoplasmic tail is also related to that of CD33, containing two tyrosine residues embodied in immunoreceptor tyrosine-based inhibitory motif-like motifs. The siglec-5 gene was shown to map to chromosome 19q13.41-43, closely linked to the CD33 gene. When siglec-5 was expressed on COS cells or as a recombinant protein fused to the Fc region of human IgG1, it was able to mediate sialic acid-dependent binding to human erythrocytes and soluble glycoconjugates, suggesting that it may be involved in cell-cell interactions. By using specific antibodies, siglec-5 was found to have an expression pattern distinct from that of CD33, being present at relatively high levels on neutrophils but absent from leukemic cell lines representing early stages of myelomonocytic differentiation. Western blot analysis of neutrophil lysates indicated that siglec-5 exists as a disulfide-linked dimer of approximately 140 kD.

Amino Acid Sequence↗
PubMed1998

Rheumatoid factor specificity of a VH3-encoded antibody is dependent on the heavy chain CDR3 region and is independent of protein A binding.

Rheumatoid factors (RF) recognize conformational determinants located within the Fc portion of IgG. By analyzing a panel of monoclonal rheumatoid arthritis (RA)-derived RFs, we previously demonstrated that the somatically generated light chain complementarity-determining region 3 (CDR3) contributes to RF specificity. We have now generated a panel of heavy chain mutants of the B'20 Ab, a high affinity RA-derived IgM RF. B'20 also binds avidly to protein A and weakly to ssDNA and tetanus toxoid. B9601, a RF negative Ab that is highly homologous to B'20 but does not bind any of the Ags tested, and RC1, a low affinity polyreactive RF, were used to generate heavy chain mutants with framework (FR) and CDR switches. The mutated heavy chains were cotransfected into a myeloma cell line with the germline counterpart of the B'20 light chain, and the expressed Ig tested for antigenic specificity. We show that both RF specificity and polyreactivity of B'20 is dependent on its unique heavy chain CDR3 region. Replacement with a B9601 CDR3 shortened to the same length as the B'20 CDR3, and with only 5 amino acid differences, did not restore Fc binding. Conversely, absence of protein A binding of B9601 is due to the presence of a serine residue at position 82a in the B9601 heavy chain FR3 region. Together, our data suggest that Ig gene recombination events can generate B cells with autoantibody specificities in the preimmune repertoire. Abnormal release, activation, expansion, or mutation of such cells might all contribute to the generation of a high titer RF response in patients with RA.

Amino Acid Sequence↗
PubMed1998

Crystal structure of a human low molecular weight phosphotyrosyl phosphatase. Implications for substrate specificity.

The low molecular weight phosphotyrosine phosphatases (PTPases) constitute a distinctive class of phosphotyrosine phosphatases that is widely distributed among vertebrate and invertebrate organisms. In vertebrates, two isoenzymes of these low molecular weight PTPases are commonly expressed. The two human isoenzymes, HCPTPA and HCPTPB, differ in an alternatively spliced sequence (residues 40-73) referred to as the variable loop, resulting in isoenzymes that have different substrate specificities and inhibitor/activator responses. We present here the x-ray crystallographic structure of a human low molecular weight PTPase solved by molecular replacement to 2.2 A. The structure of human low molecular weight PTPase is compared with a structure representing the other isoenzyme in this PTPase class, in each case with a sulfonate inhibitor bound to the active site. Possible aromatic residue interactions with the phosphotyrosine substrate are proposed from an examination of the binding site of the inhibitors. Differences are observed in the variable loop region, which forms one wall and the floor of a long crevice leading from the active-site loop. A set of residues lying along this crevice (amino acids 49, 50, and 53) is suggested to be responsible for differences in substrate specificity in these two enzymes.

Alternative Splicing↗
PubMed1998

Dendritic cell heterogeneity in vivo: two functionally different dendritic cell populations in rat intestinal lymph can be distinguished by CD4 expression.

DC derived from rat pseudo-afferent lymph (L-DC) vary in CD4, CD11b/c, Thy1, and OX41 expression. CD4 and OX41 are expressed by the same subpopulation (50-60%) of L-DC. CD4+/OX41+ L-DC express short fine processes and low nonspecific esterase, whereas CD4- DC/OX41- express long pseudopodia, high nonspecific esterase, and many cytoplasmic inclusions. These differences are stable in culture. Both populations express similar amounts of MHC class II, ICAM-1, CD11b/c and OX62. Most CD4-/OX41- L-DC are strongly positive for B7, but CD4+ L-DC express less B7, and some may be negative. Both populations express invariant chain, but both the absolute numbers and levels of expression were higher for CD4- DC. Surprisingly, CD4+ L-DC are more potent APC than CD4- cells in MLRs, for sensitized T cells in vitro and for naive T cells in vivo. Cultured CD4+/OX41+ DC can still process and present native Ag. Cultured CD4-/OX41- cells cannot present native Ag but can stimulate strong MLRs. CD4- DC invariant chain expression decreases in culture, whereas expression by CD4+ DC is stable for 48 h. CD4+ and CD4- L-DC have similar turnover times in vivo, suggesting that one population is not the precursor of the other. Thus, two separate DC populations that differ functionally and phenotypically migrate from intestine to mesenteric nodes. This may reflect distinct DC lineages or differentiation modulated by different microenvironmental stimuli.

Animals↗
PubMed1998

Activated RSC-nucleosome complex and persistently altered form of the nucleosome.

RSC, an abundant, essential chromatin-remodeling complex, related to SWI/SNF complex, binds nucleosomes and naked DNA with comparable affinities, as shown by gel shift analysis. The RSC-nucleosome complex is converted in the presence of ATP to a slower migrating form. This activated complex exhibits greatly increased susceptibility to endo- and exonucleases but retains a full complement of histones. Activation persists in the absence of ATP, and on removal of RSC, the nucleosome is released in an altered form, with a diminished electrophoretic mobility, greater sedimentation rate, and marked instability at elevated ionic strength. The reaction is reversible in the presence of RSC and ATP, with conversion of the altered form back to the nucleosome.

Adenosine Triphosphate↗
PubMed1998

Identification of functional exonic splicing enhancer motifs recognized by individual SR proteins.

Using an in vitro randomization and functional selection procedure, we have identified three novel classes of exonic splicing enhancers (ESEs) recognized by human SF2/ASF, SRp40, and SRp55, respectively. These ESEs are functional in splicing and are highly specific. For SF2/ASF and SRp55, in most cases, only the cognate SR protein can efficiently recognize an ESE and activate splicing. In contrast, the SRp40-selected ESEs can function with either SRp40 or SRp55, but not with SF2/ASF. UV cross-linking/competition and immunoprecipitation experiments showed that SR proteins recognize their cognate ESEs in nuclear extract by direct and specific binding. A motif search algorithm was used to derive consensus sequences for ESEs recognized by these SR proteins. Each SR protein yielded a distinct 5- to 7-nucleotide degenerate consensus. These three consensus sequences occur at higher frequencies in exons than in introns and may thus help define exon-intron boundaries. They occur in clusters within regions corresponding to naturally occurring, mapped ESEs. We conclude that a remarkably diverse set of sequences can function as ESEs. The degeneracy of these motifs is consistent with the fact that exonic enhancers evolved within extremely diverse protein coding sequences and are recognized by a small number of SR proteins that bind RNA with limited sequence specificity.

Animals↗
PubMed1998

OFQ reverses the kappa-opioid receptor-mediated depression of calcium current in rat dorsal root ganglion neurons.

Since the characterization of orphanin FQ (OFQ), the endogenous ligand of ORL1 receptor, much work has focused on its physiological functions. OFQ was reported to antagonize the effect of opioid-induced antinociception, although its mechanism remains obscure. In the present study, whole-cell patch clamp recording technique was used to observe if OFQ can reverse the inhibition of calcium current produced by the kappa-opioid agonist U50,488H (U50) in acutely dissociated rat DRG neurons. The concentrations of OFQ and U50 were 50 nM and 10 microM, respectively. Among 49 cells recorded, the calcium channel currents of 37 (75.5%) cells were inhibited by U50, of which 30 (81.1%) cells could be reversed by OFQ. It was interesting to note the similarity between OFQ and the well characterized anti-opioid peptide CCK-8 in that it reversed kappa-opioid receptor agonist induced suppression on calcium channel current, while by itself showed a calcium channel suppressive effect. Thus OFQ may be regarded as another anti-opioid peptide.

Animals↗