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Biomedical subjects

D Zhu

Publications and source records attributed to D Zhu.

At least 217 records · Page 12Linked to original sources
PubMed1996

Biological activities of human tumor necrosis factor-alpha and its novel mutants.

Biological activities of human tumor necrosis factor-alpha (hTNF-alpha) and its mutants were compared. In cytotoxicity assay with L929 cells, one mutant, designated as TNF-B, showed 4.5-fold higher activity than TNF examined. In receptor binding assay, TNF-B had almost the same affinity for TNF receptors on L929 cells as hTNF-alpha. We also found that TNF-B retained the cytotoxicity of hTNF-alpha for HEp-2 cells. TNF-B also had two-fold higher affinity than hTNF-alpha for receptors on HEp-2 cells (only carrying hTNF-R55) and lower affinity for receptors on U937 cells (expressing mainly hTNF-R75). These results suggested that TNF-B might still interact with the human TNF-R55 receptor, but it might largely lose its ability to bind to human TNF-R75. Changes of biological activity of TNFs might be due to an altered affinity to the different types of TNF receptor on the target cells.

Amino Acid Sequence↗
PubMed1996

13C-methacetin breath test parameter S for liver diseases diagnosis.

The mechanism of 13C-methacetin breath test is set forth clearly with the analysis of pharmacokinetics mode, and the measuring method of 13C-methacetin breath test and its clinical applications in the diagnosis of liver diseases are reported in detail. On the basis of comprehensive analysis of the clinical test data, the advanced diagnostic parameter S is of important significance for the application and development of breath test.

Acetamides↗
PubMed1996

Calcineurin mutants render T lymphocytes resistant to cyclosporin A.

The immunosuppressants cyclosporin A (CsA) and FK506 have been widely used to prevent and treat graft rejection after human organ and tissue transplantations. CsA and FK506 associate with intracellular binding proteins (i.e., CsA with cyclophilin A and FK506 with FKBP12) to form protein/drug complexes that suppress the immune system by preventing activation of T cells in response to antigen presentation. The common target of CsA and FK506 is calcineurin, a Ca2+/calmodulin-regulated, serine/threonine-specific protein phosphatase that regulates the nuclear import of a transcription factor, NF-AT, required for expression of T cell activation genes. In previous studies, we identified calcineurin mutations that block binding by the cyclophilin A/CsA or FKBP12/FK506 complexes and thereby render yeast cells resistant to the antifungal effects of CsA or FK506. In this report, we demonstrate that the corresponding mutations in murine calcineurin render the T cell receptor signal transduction cascade CsA resistant in human Jurkat T cells. Our findings support the recently determined calcineurin X-ray crystal structure, provide evidence that calcineurin is the only CsA-sensitive component limiting signaling from the T cell receptor to the nucleus, and suggest a means to render cells and tissues resistant to the toxic side effects of CsA and FK506.

Amino Acid Sequence↗
PubMed1996

Expression of biologically active human granulocyte-macrophage colony-stimulating factor in the silkworm (Bombyx mori).

Using virus derived from Bombyx mori (silkworm) nuclear polyhedrosis virus (BmNPV), we constructed an infectious recombinant virus carrying the human granulocyte-macrophage colony-stimulating factor (hGM-CSF) cDNA placed downstream from the polyhedrin promoter. The BmN cells infected with the recombinant baculovirus BmNPV-GM-CSF expressed recombinant (r)hGM-CSF up to 3.9 x 10(5) colony-formation units/ml in the medium. The maximum activity of rhGM-CSF was 4.3 x 10(6) colony-formation units/ml in the haemolymph of silkworm (Bombyx mori) larvae infected with BmNPV-GM-CSF. Three distinct species of GM-CSF of molecular masses 15, 18 and 20 kDa were detected by immunoblotting. The specific activity of rhGM-CSF purified from the haemolymph was up to 2.92 x 10(7) colony-formation units/mg. The isoelectric point of the purified rhGM-CSF was 5.5-5.7.

Animals↗
PubMed1996

[The changes in firing rates of ventromedial hypothalamic neurons and its modulations by electroacupuncture, arcuate nucleus and locus coeruleus during the immune response].

30 adult SD rats were divided into 3 grous: normal control goup; SRBC immune group (on 5th day after SRBC injection) and MSG(MSG injection at newborn stage) + SRBC group. The changes of the spontaneous firing rates of ventromedial hypothalamus (VMH) neurons and evoked responses by electroacupuncture (EA) and locus coeruleus (LC) stimulation were recorded. The results were as follows: (1) The spontaneous firing rate of VMH neurons was significant increase in the SRBC group. The firing rate in the MSG + SRBC group was higher than that in the control group, and lower than that in the SRBC group. There were statistical differences between them. (2) EA can increase the firing rates in all groups. The effect of EA was the most significant in the SRBC group, and more significant in the MSG + SRBC group. (3) The firing type of most VMH neurons showed first excited then inhibited biphasic response following stimulation LC in the SRBC and the MSG + SRBC groups. EA showed some increase in effect of the excited modulation from LC to VMH in the SRBC group. The results suggest that VMH might be an important neuroendocrinal link in immunoresponse. EA can increase the immune function of SRBC on VMH. LC and beta-endorphine neurons of arcuate nucleus might participate in the EA mediated immunoresponse.

Animals↗
PubMed1996

Urokinase mutant with better fibrin-specificity.

A 150-156 amino acids-deleted single-chain urokinase-type plasminogen activator (dscu-PA) and its recombinant wild-type counterpart (rscu-PA) were both expressed in Escherichia coli. After denaturation and renaturation in vitro, the expressed products were both purified to a single silver-stained band by means of IgG affinity chromatography. After activation by plasmin, similar enzymatic constants based on the hydrolysis of synthetic substrate S2444 by the two-chain molecular forms of dscu-PA and rscu-PA, or native tcu-PA were observed, suggesting that no impairment had been exerted on the catalytic active site of dtcu-PA by the 150-156 amino acids deletion. In both in vitro fibrin-clot and 125I-fibrin sepharose lysis tests, dtcu-PA showed a significantly higher fibrinolytic activity than rtcu-PA or rscu-PA. Hardly any effect on the concentration of fibrinogen in plasma was found in dtcu-PA. It was concluded that dtcu-PA had a higher fibrin specificity and that tcu-PA could be provided with better fibrin specificity by means of mutation.

Amino Acid Sequence↗
PubMed1995

A genetic approach to idiotypic vaccination for B cell lymphoma.

Idiotypic immunoglobulin expressed by a B cell tumor presents a clear tumor antigen which could be attacked by vaccination of the host. Vaccination with idiotypic protein has been shown to induce protective immunity against lymphoma, but application to patients is limited by the requirement of "personal" vaccines for each patient. A genetic approach enables V-region sequences encoding idiotypic antigen to be rescued from tumor biopsies, and to be assembled as scFv fragments. These can be expressed in bacteria to produce recombinant protein, or used directly as naked DNA vaccines. Intramuscular injection of idiotypic DNA from a mouse B cell lymphoma induces low levels of syngeneic anti-idiotypic antibody in serum. Response can be stimulated by co-injection of DNA plasmids encoding either IL-2 or GM-CSF, and T cells which proliferate in response to idiotypic IgM are generated. However, protection against tumor appears to be blocked by continuing secretion of idiotypic antigen from the persisting vaccine vector, which forms immune complexes with serum antibody. Methods for regulating the level of scFv to engage the immune system, but not to block the effector arm are being investigated. Similar control will be applicable to the cytokine vectors, which can deliver encoded cytokines designed to activate immune pathways for tumor destruction. Experience gained in lymphoma may be extended to other tumors with defined tumor antigens.

Adjuvants, Immunologic↗
PubMed1995

Myristoylation of calcineurin B is not required for function or interaction with immunophilin-immunosuppressant complexes in the yeast Saccharomyces cerevisiae.

Calcineurin is a heterodimeric Ca2+/calmodulin-dependent protein phosphatase that regulates signal transduction and is the target of immunophilin-immunosuppressive drug complexes in T-lymphocytes and in yeast. Calcineurin is composed of a catalytic A subunit and a regulatory B subunit that is myristoylated at its amino terminus. We employed genetic and biochemical approaches to investigate the functional roles of myristoylation of calcineurin B (CNB1) in Saccharomyces cerevisiae. A calcineurin B mutant in which glycine 2 was substituted by alanine (CNB1-G2A) did not incorporate [3H]myristate when expressed in yeast. Both wild-type calcineurin B and the CNB1-G2A mutant protein are partially associated with membranes and cytoskeletal structures; hence, myristoylation is not required for these associations. In several independent genetic assays of calcineurin functions (recovery from alpha-factor arrest, survival during cation stress, and viability of a calcineurin-dependent strain), the nonmyristoylated CNB1-G2A mutant protein exhibited full biological activity. In vitro, both wild-type and CNB1-G2A mutant proteins formed complexes with both cyclophilin A-cyclosporin A (CsA) and FKBP12-FK506 that contained calcineurin A. Interestingly, expression of the nonmyristoylated CNB1-G2A mutant protein rendered yeast cells partially resistant to the immunosuppressant CsA, but not to FK506. This study demonstrates that calcineurin B myristoylation is not required for function, but may participate in inhibition by the cyclophilin A-CsA complex.

Amino Acid Isomerases↗
PubMed1995

Cardiac malformation in neonatal mice lacking connexin43.

Gap junctions are made up of connexin proteins, which comprise a multigene family in mammals. Targeted mutagenesis of connexin43 (Cx43), one of the most prevalent connexin proteins, showed that its absence was compatible with survival of mouse embryos to term, even though mutant cell lines showed reduced dye coupling in vitro. However, mutant embryos died at birth, as a result of a failure in pulmonary gas exchange caused by a swelling and blockage of the right ventricular outflow tract from the heart. This finding suggests that Cx43 plays an essential role in heart development but that there is functional compensation among connexins in other parts of the developing fetus.

Animals↗
PubMed1995

Splenic lymphoma with villous lymphocytes involves B cells with extensively mutated Ig heavy chain variable region genes.

Splenic lymphoma with villous lymphocytes (SLVL) is a recently defined subgroup of chronic B-cell lymphoproliferative diseases. The characteristic morphology of the tumor cells, together with phenotypic and cytogenetic findings, indicate that it is a distinct entity, but the nature of the cell or origin and its relationship to other low-grade lymphomas is unclear. For B-cell tumors, analysis of the variable region heavy chain (VH) genes used to encode the clonal Ig has shown marked differences between histologic categories, both in gene usage and extent of somatic mutation. An investigation of VH genes used in five typical cases of SLVL has shown somatic hypermutation from germline sequences in all cases, indicating that the cell of origin has been exposed to the hypermutation mechanism. However, no clonal heterogeneity was detectable, demonstrating that the tumor cell does not accumulate further mutations. These characteristics are similar to those found in mature postfollicular B cells, such as plasma cells. The distribution of mutations leading to replacement amino acids differed among the cases, with three of five cases showing clear evidence for antigen selection.

Aged↗
PubMed1995

Association and linkage with quantitative traits.

This paper examines two approaches for the analysis of quantitative traits: (1) association studies and (2) linkage studies. The trait studied was Q1 from simulated Problem 2 data set in Genetic Analysis Workshop 9. Our purpose was to evaluate associations present in the data, to identify nongenetic and genetic predictors of the trait, and to explore the simulated genome for linkage. Through the association study, we found evidence for the primary major gene associated with this trait. The linkage study found evidence of residual genetic effect acting through other traits. Adjustments of Q1 for Q2 and Q3 led to a failure to find significant effects of MG2 and MG3. This supports the suggestion that adjustment for genetically influenced traits for effects of other genetic traits can reduce the power to detect major gene effects. In summary, we detected the major gene directly associated with the trait of interest through association studies. Linkage analysis detected evidence for two other genes associated to a lesser degree with the trait.

Analysis of Variance↗
PubMed1995

Interaction of CD44 variant isoforms with hyaluronic acid and the cytoskeleton in human prostate cancer cells.

CD44 is a glycosylated adhesion molecule which may undergo alternative splicing of 10 possible exons to generate variant isoforms. A number of CD44 variant isoforms expressed by tumor cells have been correlated with metastatic and proliferative behavior. In this study, we have characterized CD44 isoform expression on three prostate cancer cell lines: ALVA-31, PPC-1, and LNCaP. Using reverse transcriptase-polymerase chain reaction, we have found that ALVA-31 and PPC-1 cells express multiple CD44 isoforms, including CD44s (standard form), CD44E (epithelial form), and an exon 14-containing form. In addition, two smaller forms have been detected: one using an alternative donor splice site within exon 5, and a novel form omitting exon 5 entirely. The CD44 isoforms expressed by ALVA-31 and PPC-1 cells appear to be preferentially located on the cell surface. By contrast, LNCaP cells do not express any of the CD44 forms at the RNA or protein level. Both PPC-1 and ALVA-31 cells display tumorigenesis and invasiveness in nude mice, whereas LNCap cells exhibit a less malignant phenotype, suggesting a correlation between CD44 variant (CD44v) expression and aggressive prostate tumor behavior. Functional characterization reveals that CD44 mediates prostate cell adhesion to extracellular hyaluronic acid (HA). In addition, the CD44 cytoplasmic domain binds specifically to ankyrin, a membrane cytoskeletal protein. Double immunofluorescence labeling and confocal microscopic analyses indicate that HA binding induces the HA receptor (i.e., CD44) to form capped structures. Importantly, intracellular ankyrin is preferentially accumulated underneath HA receptor-capped structures. These results suggest that cytoskeletal proteins such as ankyrin are closely associated with CD44-mediated signaling events induced by HA. Finally, HA-mediated transmembrane interactions between CD44 isoforms and cytoskeletal proteins (i.e. ankyrin) may play a pivotal role in regulating tumor cell behavior during human prostate cancer development.

Ankyrins↗
PubMed1995

Involvement of CD44 and its variant isoforms in membrane-cytoskeleton interaction, cell adhesion and tumor metastasis.

CD44s (standard form of CD44) is a transmembrane glycoprotein whose external domain displays extracellular matrix adhesion properties by binding both hyaluronic acid (HA) and collagen. The cytoplasmic domain of CD44s interacts with the cytoskeleton by binding directly to ankyrin. It has been shown that post-translational modifications, such as phosphorylation (by protein kinase C), acylation (by acyl-transferase) and GTP-binding enhanced CD44's interaction with cytoskeletal proteins. Most importantly, the interaction between CD44s and the cytoskeletal protein, ankyrin, is required for the modulation of CD44s cell surface expression and its adhesion function. Recently, a number of tumor cells and tissues have been shown to express CD44 variant (CD44v) isoforms. Using RT-PCR and DNA sequence analyses, we have found that unique CD44 splice variant isoforms are expressed in both prostate and breast cancer cell lines and carcinomas. Most importantly intracellular ankyrin is preferentially accumulated underneath the patched/capped structures of CD44 variant isoform in both breast and prostate cancer cells attached to HA-coated plates. We propose that selective expression of CD44v isoforms unique for certain metastatic carcinomas and their interaction with the cytoskeleton may play a pivotal role in regulating tumor cell behavior during tumor development and metastasis.

Animals↗
PubMed1995

Centromere spreading and centromeric aberrations in ovarian tumors.

Centromere spreading (CS) of chromosomes and high occurrence of aberrations at centromeric region were observed in two papillary serous cystadenocarcinomas and one borderline papillary serous cystadenoma of the ovary. In the borderline tumor, CS of chromosome 12, trisomy of which had been reported as the sole abnomaly in benign ovarian tumors, was seen in three metaphases. It is suggested that CS may be an early event in the carcinogenesis of epithelial ovarian tumors. Centromeres of some chromosomes in tumor cells may be unstable and thus bring about premature centromere separations and breakages at centromeric regions, followed by some numerical and structural chromosomal aberrations.

Centromere↗
PubMed1995

The maize mitochondrial MnSODS encoded by multiple genes are localized in the mitochondrial matrix of transformed yeast cells.

We recently demonstrated that maize mitochondrial MnSODs are encoded by a differentially expressed multigene family. Molecular characterization of the cloned maize MnSod cDNAs shows that all the maize MnSODs encoded by the cloned MnSod have mitochondrial transit peptides and that the matrix-targeting signal was conserved in all the MnSOD precursor proteins, suggesting that all the maize MnSODs are associated with mitochondria in vivo. In an attempt to clarify the submitochondrial localization of each individual maize MnSOD, each of the maize MnSod cDNAs was introduced into the MnSOD-deficient yeast cells. The functional role and submitochondrial location of each individual MnSOD were examined in the transformed yeast cells. Herein, we demonstrate that all the maize MnSODs can functionally complement the MnSOD deficiency and rescue the transformed yeast cells from oxidative stress induced by paraquat. Biochemical analyses indicate that all of the maize MnSODs are expressed and targeted into the mitochondrial matrix of transformed yeast cells.

Gene Expression↗
PubMed1995

Molecular mechanisms of immunosuppression by cyclosporine, FK506, and rapamycin.

The immunosuppressant cyclosporine A revolutionized treatment of graft rejection. Two newer agents, FK506 and rapamycin, show great clinical potential. These drugs suppress the immune system by forming protein-drug complexes that interact with and inhibit key components of the signal transduction pathways required for T-cell activation. The target of the cyclophilin A-cyclosporine A and FKBP12-FK506 complexes is calcineurin, a protein phosphatase required for signaling via the T-cell receptor. Cyclosporine A and FK506 nephrotoxicity may reflect renal-specific functions of calcineurin. The target of the FKBP12-rapamycin complex is TOR, a lipid and protein kinase homolog that is likely to be required for T-cell proliferation in response to interleukin-2. The identification of cyclosporine A, FK506, and rapamycin targets reveals much concerning T-cell signaling and provides the means to design novel immunosuppressants with reduced toxicity.

Animals↗