Biomedical subjects

Z Liu

Publications and source records attributed to Z Liu.

At least 1,153 records · Page 64Linked to original sources

PubMed1994

Effect of acute diabetes on rat hepatic glucose-6-phosphatase activity and its messenger RNA level.

Glucose-6-phosphatase catalyzes the final step of glucose production by liver and kidney. Though its strategic position has sparked interest in its regulation, difficulty with isolating a pure, stable enzyme has slowed progress. Virtually all previous work examining the physiologic regulation of this enzyme has relied on estimates of glucose-6-phosphatase activity in crude microsome preparations. The recent cloning of human and murine glucose-6-phosphatase cDNAs has now allowed study of its mRNA expression. We studied the effect of acute, streptozotocin-induced diabetes on hepatic microsomal glucose-6-phosphatase activity and mRNA expression in young (89 +/- 3 g), juvenile (304 +/- 4 g) and adult (512 +/- 10 g) rats. In control rats, mRNA expression and enzyme activity was similar among the three age groups. Streptozotocin-induced diabetes significantly increased the enzyme activities in both intact and triton-treated microsomes in all groups of rats (p < 0.001). Glucose-6-phosphatase mRNA expression was increased in the diabetic rats as well (p < 0.0001). Blood glucose concentrations correlated significantly with glucose-6-phosphatase mRNA level (p < 0.005) and both intact (p < 0.002) and triton-treated (p < 0.001) microsomal glucose-6-phosphatase activity. Both intact and triton-treated microsomal glucose-6-phosphatase activity correlated with mRNA level (p < 0.001, for each). We conclude that acute streptozotocin-diabetes increase expression of glucose-6-phosphatase mRNA and this contributes to the increased glucose-6-phosphatase activity seen with diabetes mellitus.

Acute Disease↗

PubMed1994

Beta-structure in human amylin and two designer beta-peptides: CD and NMR spectroscopic comparisons suggest soluble beta-oligomers and the absence of significant populations of beta-strand dimers.

Intensity variation for the positive far UV CD band was observed for three 'beta-sheet' peptides. In 6% HFIP, an amyloidogenic species (human pancreatic amylin) displays, on standing, an extremely intense 192-nm band which diminishes upon physical agitation. A concurrently formed Tyr sidechain band at 274 nm disappears completely with agitation, linking the enhancement of the 192-nm band to the highly ordered stacking of beta-sheets. NMR studies indicate that the beta-states of the three peptides are oligomeric, not beta dimers. A membrane-forming EAK peptide displays NMR peaks due to the low concentration of 'random coil' monomers present in slow equilibrium with beta-oligomers; solutions of a more hydrophobic ELKA peptide, which displays an intense 195-nm band, contain only oligomeric species. NMR studies at 25% HFIP revealed the structural requirements for inhibition of beta-oligomer formation.

Amino Acid Sequence↗

PubMed1994

Expression of vesicle-associated membrane protein 2 (VAMP-2)/synaptobrevin II and cellubrevin in rat skeletal muscle and in a muscle cell line.

Molecular studies have identified a family of synaptic vesicle-associated membrane proteins (VAMPs, also known as synaptobrevins) which have been implicated in synaptic vesicle docking and/or fusion with plasma membrane proteins. Here we demonstrate the expression of two members of this family, VAMP-2/synaptobrevin II and cellubrevin, in skeletal muscle, a tissue with both constitutive and regulated membrane traffic. The 18 kDa VAMP-2 polypeptide was detected in purified membrane fractions from adult skeletal muscle and from L6 myotubes in culture, demonstrating that the presence of this protein in the isolated muscle membrane fractions is not the result of contamination by ancillary tissues such as peripheral nerve. Furthermore, skeletal muscle and the muscle cell line also expressed cellubrevin, a VAMP-2 homologue of 17 kDa; which is much less abundant in brain cells. Both VAMP-2 and cellubrevin were preferentially isolated in membrane fractions rich in plasma membranes, and were less concentrated in light microsomes and other internal membrane fractions of mature muscle or muscle cells in culture. Interestingly, both VAMP-2 and cellubrevin were much more abundant in the differentiated L6 myotubes than in their precursor myoblasts, suggesting that they are required for functions of differentiated muscle cells. The identity of both polypeptides was further confirmed by their susceptibility to proteolysis by Clostridium tetanus toxin. Expression of these products was further established by the presence of mRNA transcripts of VAMP-2 and cellubrevin, but not of VAMP-1, in both skeletal muscle and L6 myotubes. In contrast, other synaptic vesicle and docking/fusion components were undetectable, such as VAMP-1, SNAP25 and syntaxin 1A/1B, as were synaptophysin and synapsin Ia/Ib, proteins which are believed to be involved in sensing the signal for neuronal exocytosis. It is concluded that VAMP-2 and cellubrevin are expressed in skeletal muscle cells and may each participate in specific processes of intracellular membrane traffic.

Amino Acid Sequence↗

PubMed1994

Resolution of proteins on a phenyl-Superose HR5/5 column and its application to examining the conformation homogeneity of refolded recombinant staphylococcal nuclease.

In order to examine the effect of amino acid substitutions on protein retention in hydrophobic interaction chromatography and the resolution of a phenyl-Superose HR5/5 column, two groups of staphylococcal nucleases, named Y113/W140 (wild-type), Y113W/W140 and Y113/W140F, Y113W/W140F, were produced by substituting tryptophan (W) for tyrosine (Y) at residue 113 and phenylalanine (F) for tryptophan (W) at residue 140. For each group, the proteins have the same amino acid at residue 140, but a different amino acid at residue 113. The solvent perturbation of nuclease fluorescence and 1,8-anilinoaphthalene-8-sulfonate binding studies showed that the substitutions do not change the side-chain positions of amino acids at residues 113 and 140. Chromatography of the proteins on the Phenyl-Superose HR5/5 column showed that the proteins with tryptophan at residue 113 have longer retention times than the proteins having tyrosine at residue 113; the proteins with the same amino acid at residue 113 have almost the same retention time regardless of substituting phenylalanine for tryptophan at residue 140. The studies clearly indicate that not all amino acid substitutions have an effect on protein retention; the contribution to retention of a given amino acid substitution depends on its position in a protein. Single amino acid substitutions at the exterior surface of a protein, which change the strength of hydrophobic interaction, can affect the protein retention in hydrophobic interaction chromatography. Staphylococcal nuclease and its mutants with only one amino acid difference on their surfaces can be discriminated by the phenyl-Superose column.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗

PubMed1994

Identification of functional cysteine residues in human galactosyltransferase.

The functions of the five cysteine residues in human galactosyltransferase were investigated using site-directed mutagenesis to determine the location of the disulfide bond as well as the role of the sulfhydryl groups. The enzyme remains active when three of its cysteine residues at positions 171, 264 and 340 are mutated to serine separately. However, enzymatic activity is lost when either cysteine-129 or cysteine-245 is replaced with serine. The loss of GT activity suggests that these two cysteine residues form a disulfide bond. The three active mutated enzymes were studied kinetically. The kinetic constants of the enzymes with cysteine-171 or cysteine-264 replaced with serine are not significantly different from those of GT that does not have these substitutions. When cysteine-340 was mutated, however, the kinetic constant for UDP-galactose increased about 30 fold, while that for N-acetylglucosamine and Mn2+ remained unchanged. In addition, sulfhydryl inhibition studies reveal that cysteine-340 is the only cysteine residue that reacts with the sulfhydryl reagents. These results indicate that cysteine-340 may be involved in the binding of UDP-galactose.

Cysteine↗

PubMed1994

Simultaneous activation of Ig and Oct-2 synthesis and reduction of surface MHC class II expression by IL-6.

Terminal differentiation of B cells to plasma cells in vivo is characterized by secretion of Ig and extinction of MHC class II expression on the cell surface. We show that IL-6 signaling leads to marked increases in the synthesis and secretion of Ig in clonal human B cell lines and newly isolated polyclonal B lymphocytes in vitro. The IL-6-induced cells resemble plasma cells in ultrastructure and in reduced expression of surface MHC class II. Enhanced Ig synthesis is a result of coordinated transcriptional activation of Ig genes without promoter or isotype specificity, and differential accumulation of the mRNA encoding the secreted form of Ig heavy chain. It is saturable and subject to negative control when IL-6 stimulation is prolonged. Coordinate with temporal changes in Ig synthesis, the DNA-binding activity and the synthesis of the B cell-enriched transcription factor Oct-2 are regulated. Thus, differentiation of B cells with IL-6 in vitro recapitulates the hallmarks of terminal B differentiation in vivo; Oct-2 may have a role in this process.

Antibody Formation↗

PubMed1994

Impairment of place navigation of rats in the Morris water maze by intermittent light is inversely related to the duration of the flash.

The relative contribution of allocentric and egocentric orientation to place navigation was studied in Long-Evans rats trained in the Morris water maze in permanent light, permanent darkness or flickering light (1 Hz, flash durations 25, 100, 300, 500 and 800 ms). After 3 days of training (nine blocks of four trials), escape latencies were 38 and 7 s in the dark- and light-trained groups, respectively, and corresponded to the light-dark ratio in the flicker-trained groups. Shorter-than-predicted latencies in the 25- and 100-ms groups reflected visual persistence of approximately 200 ms. The difference between flickering light (100 ms) and permanent light performance during acquisition of place navigation to a new target was significantly smaller in rats previously trained in light than in naive animals. It is concluded that longer flash duration gives the animals more opportunities to locate relevant landmarks and to estimate their distance.

Animals↗

PubMed1994

Comprehensive two-dimensional gas chromatography for the fast separation and determination of pesticides extracted from human serum.

A comprehensive two-dimensional gas chromatograph with flame ionization detection was constructed and evaluated for the fast separation and analysis of pesticides. A two-stage thermal desorption modulator served as an interface between the two capillary GC columns. By controlling the temperature of the modulator chamber, all sample substances covering a wide polarity and volatility range were modulated without sample breakthrough. Orthogonal separations were achieved with a nonpolar first column and a moderately polar second column. The system allows fast separations of complex mixtures. When we used the method to analyze pesticides extracted from human samples, we achieved complete separation of 15 pesticides in less than 4 min. The flame ionization detector gave detection limits for particular pesticides that ranged from 1.8 to 3.8 pg on-column. The relative standard deviations were from 6.2% to 8.8% over the linear dynamic range.

Chromatography, Gas↗

PubMed1994

Some physicochemical and enzymic properties of selenium-containing abzyme.

We successfully prepared the Se-containing abzyme (Se-abzyme) with glutathione peroxidase (GPX) activity and further studied its physicochemical and enzymic properties and stabilities. Data showed that the isoelectric point of the abzyme was 6.95-7.08, and its molecular weight was 158 KD. The ranges of optimum pH and temperature of the Se-abzyme were wider than the native GPX. The store stability of the abzyme was higher than the native GPX. The Se content in the abzyme was found to be 5 mol Se/mol abzyme by X-ray photoelectron spectrum, and binding constant 1.11 x 10(7)M-1 by using ELISA method. The Se-abzyme was inhibited competitively by dithiobis(2-nitrobenzoic acid) (DTNB), and inhibition constant was determined to be 1.25 x 10(-3)M-1.

Electrophoresis, Polyacrylamide Gel↗

PubMed1994

Effects of cerebellar lesions on tonic seizures, tremor and lifespan in myelin-deficient rats.

In common with other dysmyelinating mutants, the myelin-deficient rat displays an action tremor and tonic seizures culminating in the death of the animals at approximately 23-26 days. We find that deep lesions of the cerebellar vermis alleviate the manifestations of the myelin deficiency significantly. Such lesions introduced at 20 days or later eliminate both tremor and seizures for periods up to 10 days. Lifespan is prolonged to nearly 30 days, on average, and to 35 days in some cases. Shallow lesions of the vermis or lateral lobe lesions have relatively little effect. Based on these observations we suggest that the cerebellum contributes not only to the action tremor but also to the tonic seizures characteristic of central myelin deficiency. Spontaneous activity originating in myelin-deficient fiber tracts may be carried to the cerebellum and processed there to produce a highly amplified and/or synchronized output to broad areas of the neuraxis. Deep lesions of the vermis presumably interfere with cerebellar output and compromise the cerebellar contribution to the seizures. Tonic seizures and other 'paroxysmal attacks' also occur commonly in human demyelinating diseases including multiple sclerosis [11]. Manipulation of cerebellar output offers a potential approach to the control of such spontaneous activity.

Animals↗

PubMed1994

Field-amplified sample stacking in micellar electrokinetic chromatography for on-column sample concentration of neutral molecules.

On-column concentration of neutral molecules was achieved for the first time in micellar electrokinetic chromatography by means of field-amplified sample stacking. The stacking process was accomplished by dissolving the neutral analytes in a low-concentration micellar solution that was still above the critical micelle concentration. The lower total ionic strength in the sample buffer compared to the electrophoresis buffer allowed the negatively charged micelles to migrate rapidly into the boundary between the sample and the running buffer where they slow down. This field-amplified sample stacking was achieved by using normal or reversed electrode polarity and produced a 75-85-fold increase in sensitivity for 1,2,4,7- and 1,2,4,8-tetrachlorodibenzo-p-dioxins. The peak area counts obtained from the sample stacking process were proportional to the sample volume injected, and the stacking efficiency was dependent on the micellar concentration. The best stacking efficiency was obtained when the micelle concentration was slightly higher than the critical micelle concentration. When the injection volume was relatively small, the normal-polarity stacking procedure produced a higher stacking efficiency. However, when the injection volume was large, reversed polarity produced a higher stacking efficiency because the non-uniform distribution of the electrical field strength had been eliminated.

Chromatography↗

PubMed1994

The yeast KEM1 gene encodes a nuclease specific for G4 tetraplex DNA: implication of in vivo functions for this novel DNA structure.

We have previously reported the identification of a G4-DNA-dependent nuclease from S. cerevisiae that recognizes a tetrastranded G4-DNA structure and cuts in a single-stranded region 5' to the G4 structure. We purify this activity to homogeneity and show it to be the product of the S. cerevisiae KEM1 gene, which is also known as SEP1, DST2, XRN1, and RAR5. Since a homozygous deletion of the KEM1 gene blocks meiotic cells at the 4N stage, the finding of these G4-dependent DNA binding and cleavage activities for the KEM1 gene product supports the hypothesis that G4-DNA may play a role in meiosis.

Amino Acid Sequence↗

PubMed1994

Characterization of a protein that interacts with the rat ribosomal gene promoter and modulates RNA polymerase I transcription.

An RNA polymerase I core promoter binding factor (CPBF) was purified to apparent homogeneity from rat adenocarcinoma ascites cells by chromatographic fractionation on a series of columns including an oligodeoxynucleotide affinity column. The final preparation contained two polypeptides with molecular masses of 44,000 and 39,000 daltons. The binding of the factor to the promoter was demonstrated by Southwestern blotting, UV cross-linking and electrophoretic mobility shift assay. The specificity of its binding to the core promoter was confirmed by competitive electrophoretic mobility shift assay using several unlabeled oligo probes in the assay. The addition of increasing amounts of purified CPBF to the in vitro transcription reaction that contains a limiting quantity of the factor resulted in dramatic stimulation of RNA polymerase I (pol I) transcription of rat ribosomal RNA gene. The transcription stimulatory activity associated with the purified CPBF fractions co-purified with the core promoter binding activity in an electrophoretic mobility shift assay. Finally, in a reconstitution transcription system which is devoid of the factor and is incapable of ribosomal gene transcription, purified CPBF could trans-activate the pol I promoter. These data indicate that CPBF is a novel pol I promoter binding factor required for ribosomal gene transcription.

Adenocarcinoma↗

PubMed1994

Protein tyrosine phosphorylation triggered by human Fc gamma RII.

The role of the cytoplasmic domain of human Fc gamma RII in cellular activation was studied by transfecting P815 cells with different isoforms or deletion mutants of this receptor and then assaying protein tyrosine phosphorylation after crosslinking with anti-CD32 monoclonal antibodies. Fc gamma RIIa, but not Fc gamma RIIb1 or b2, was able to trigger tyrosine phosphorylation. Deletion of just the carboxyl-terminal 19 amino acids of Fc gamma RIIa markedly reduced induction of protein tyrosine phosphorylation. Deletion of the carboxyl-terminal 38 amino acids of the cytoplasmic domain completely eliminated this response. These results indicate the cytoplasmic domain of human Fc gamma RIIa, the form of Fc gamma RII that predominates on human myeloid cells and platelets, contains sequences that allow activation of tyrosine phosphorylation; Fc gamma RIIb isoforms, the form of Fc gamma RII on human B lymphocytes, lack these sequences.

Amino Acid Sequence↗

PubMed1994

Role of IL-6 in activation of T cells for acquired cellular resistance to Listeria monocytogenes.

We have previously shown that IL-6 is a major cytokine in the serum of mice infected with the intracellular bacterium Listeria monocytogenes, and that injection of rIL-6 before, but not after, infection promotes the recovery of mice from listeriosis. Here we demonstrate that IL-6 is required in the early stages of infection, in that injection of anti-IL-6 Ab 24 or 4 h before infection, but not 24 h after, led to a massive increase in bacterial numbers 4 or more days after infection. On the other hand, injection of rIL-6 before infection significantly suppressed bacterial numbers in the liver and spleen 3 to 4 days after infection and increased the production of IFN-gamma by in vitro cultured CD4+ and CD8+ T cells in response to specific Ag. rIL-6 did not protect SCID mice, which lack both T and B lymphocytes, against Listeria infection. The protective effect of rIL-6 was neutralized by the injection of Ab to IFN-gamma. We conclude that IL-6 plays an essential role in the activation of T cells to produce IFN-gamma, the cytokine that is central to acquired cellular resistance to intracellular bacteria.

Animals↗

PubMed1994

Spontaneous transformation of rat ovarian surface epithelial cells results in well to poorly differentiated tumors with a parallel range of cytogenetic complexity.

We previously used rat ovarian surface epithelial cells subjected to repetitious growth in vitro to provide experimental evidence in support of a role for incessant ovulation in the etiology of ovarian cancer. We have now initiated a series of 30 independent rat ovarian surface epithelial cell lines. This report describes findings in eight of the cell lines that, based on tumor formation in athymic mice, have undergone malignant transformation. Each of the tumors exhibited chromosomal alterations. Two well to moderately differential tumors had only one or two cytogenetic changes, and they had in common the presence of numerical gains. Each of five poorly differentiated tumors had complex karyotypes with three to eight clonal aberrations, prominent among them being unbalanced rearrangements and numerical losses. Several poorly differentiated tumors also had marker chromosomes, double minutes, or homogeneously staining regions. These findings demonstrate that the malignant tumors produced by spontaneously transformed rat ovarian surface epithelial cell lines range in degree of differentiation, which is paralleled by the cytogenetic complexity. Thus, this model system may fill an important void in future efforts to define the genetic basis of common epithelial tumors of the ovary and many features characteristic of these neoplasms.

Adenocarcinoma↗

PubMed1994

Targeted nuclear antisense RNA mimics natural antisense-induced degradation of polyoma virus early RNA.

We describe a general antisense strategy to inhibit target gene expression. The substitution of a cis-acting ribozyme for a polyadenylylation signal in an antisense expression vector results in the nuclear retention of RNAs and the efficient degradation of their targets. We demonstrate the utility of this system in polyoma virus, where early-strand RNA levels are downregulated in the nucleus by antisense late-strand counterparts. We show that mutations destabilizing these naturally occurring antisense transcripts lead to increased levels of early-strand RNAs. Furthermore, expression in trans of nuclear antisense transcripts lowers early-strand RNA levels and quantitatively mimics the natural regulation.

3T3 Cells↗

PubMed1994

Experimental studies of the inhibitory effects of green tea catechin on mice large intestinal cancers induced by 1,2-dimethylhydrazine.

Three hundred Kunming mice were randomly divided into six groups (half males and half females in each group). Group 1 was the positive control group, Groups 2, 3, 4 and 5 were experimental groups and Group 6 was used as the solvent control group. Mice in Groups 1-4 were injected with 1,2-dimethylhydrazine (1,2-DMH) (20 mg/kg body wt.) solution subcutaneously once a week from the 2nd week to the 20th week. From the 1st week to the 23rd week, mice in Groups 2, 3 and 4 were given catechin (1 mg/mouse), catechin (2 mg/mouse) and EGCG (2 mg/mouse), respectively, five times a week. Mice in Group 5 received only catechin (3 mg/mouse) five times a week from the 1st to the 23rd week. Mice in Group 6 were injected with an equal volume of 1 mmol EDTA solution subcutaneously once a week from the 2nd to the 20th week. At the end of the 27th week, all the mice were killed by cervical dislocation (Zhu, Q.H. and Zhu, Q.F. (1991) Laboratory Animal Science, 1st edition. The Junior Educational Publisher, Guangdong). Pathological examinations indicated that the incidence of large intestinal cancers occurring in Group 1 was 80%, significantly higher than that in Groups 2, 3 and 4 (p < 0.001). No tumors were found in Groups 5 and 6. This might suggest that green tea has preventive effects on large intestinal cancer induction in spite of the different doses of catechin. Immunohistochemistry studies showed that green tea catechins could enhance the activity of superoxide dismutase (SOD) in tissues.

1,2-Dimethylhydrazine↗