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Biomedical subjects

G Zhang

Publications and source records attributed to G Zhang.

At least 433 records · Page 24Linked to original sources
PubMed1997

Molecular epidemiology of recent outbreaks of swine vesicular disease: two genetically and antigenically distinct variants in Europe, 1987-94.

Viruses from the recent epidemic of swine vesicular disease (SVD) in Europe have been isolated and characterized by antigenic and genetic methods to examine the likely epidemiological origins of the disease. Antigenic analysis was performed on 77 SVD viruses (SVDV) isolated in Europe between 1966 and 1994 using two panels of monoclonal antibodies (MAb) in a trapping ELISA. Genetic analysis of 33 of the SVD viruses by reverse transcription-polymerase chain-reaction (RT-PCR) amplification and nucleotide sequencing of the ID (VP1) coding region was also performed. Comparison of the nucleotide sequences with each other and with three other previously published SVDV sequences revealed four distinct groups which correlated exactly with the results of the pattern of reactivity with MAbs. The first group consisted solely of the earliest SVD virus isolated (ITL/1/66) while the second group comprised viruses present in Europe and Japan between 1972 and 1981. The third group consisted of viruses isolated from outbreaks of SVD in Italy between December 1988 and June 1992. Viruses isolated between 1987 and 1994 from Romania, the Netherlands, Italy and Spain formed a fourth group. The genetic and antigenic similarity of the most recent virus isolates from Western Europe to a virus isolated in Romania 5 years previously suggests that the possible origin of the recent epidemic of swine vesicular disease in Western Europe was in Eastern Europe.

Animals↗
PubMed1997

Self-assembling DNA-lipid particles for gene transfer.

PURPOSE: We have demonstrated that a heteromolecular complex consisting of cationic lipids and DNA can be prepared and isolated (1). Cationic lipids bind DNA through electrostatic interactions. However, when sufficient lipids are bound to DNA the physical and chemical properties of the complex are governed by hydrophobic effects. Here we describe an approach where this hydrophobic complex is used as an intermediate in the preparation of lipid-DNA particles (LDPs). METHODS: The approach relies on the generation of mixed micelles containing the detergent, n-octyl beta-D-glucopyranoside (OGP), the cationic lipid, N-N-dioleoyl-N, N-dimethylammonium chloride (DODAC), and selected zwitterionic lipids, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) or egg sphingomyelin (SM). RESULTS: When these micelles were prepared at low detergent concentrations (20 mM OGP) and combined with pCMV beta DNA, LDPs spontaneously formed. The mean diameter of these particles as measured by quasielastic light scattering was 55-70 nm, a result that was confirmed by negative stain electron microscopy. Further characterization of these LDPs showed that DNA within the particles was inaccessible to the small fluorochrome TO-PRO-1 and protected against DNase I degradation. LDPs could also be prepared in high concentrations of OGP (100 mM), however particles formed only after removal of OGP by dialysis. Particles formed in this manner were large (> 2000 nm) and mediated efficient transfection of Chinese hamster ovary cells. Transfection activity was greater when the lipid composition used consisted of SM/ DODAC. Small particles (< 100 nm) prepared of SM/DODAC were, however, inefficient transfecting agents. CONCLUSIONS: We believe that LDP formation is a consequence of the molecular forces that promote optimal hydrocarbon-hydrocarbon interactions and elimination of the hydrocarbon-water interface.

Animals↗
PubMed1997

The relationship between protein aggregation and molecular mobility below the glass transition temperature of lyophilized formulations containing a monoclonal antibody.

PURPOSE: To find out if the physical instability of a lyophilized dosage form is related to molecular mobility below the glass transition temperature. Further, to explore if the stability data generated at temperatures below the glass transition temperature can be used to predict the stability of a lyophilized solid under recommended storage conditions. METHODS: The temperature dependence of relaxation time constant, tau, was obtained for sucrose and trehalose formulations of the monoclonal antibody (5 mg protein/vial) from enthalpy relaxation studies using differential scanning calorimetry. The non-exponentiality parameter, beta, in the relaxation behavior was also obtained using dielectric relaxation spectroscopy. RESULTS: For both sucrose and trehalose formulations, the variation in tau with temperature could be fitted Vogel-Tammann-Fulcher (VTF) equation. The two formulations exhibited difference sensitivities to temperature. Sucrose formulation was more fragile and exhibited a stronger non-Arrhenius behavior compared to trehalose formulation below glass transition. Both formulations exhibited < 2% aggregation at t/tau values < 10, where t is the time of storage. CONCLUSIONS: Since the relaxation times for sucrose and trehalose formulations at 5 degrees C are on the order of 10(8) and 10(6) hrs, it is likely that both formulations would undergo very little (< 2%) aggregation in a practical time scale under refrigerated conditions.

Antibodies, Monoclonal↗
PubMed1997

Developmentally regulated O-acetylated sialoglycans in the central nervous system revealed by a new monoclonal antibody 493D4 recognizing a wide range of O-acetylated glycoconjugates.

We have previously detected an alkali-labile and developmentally regulated antigen in rat embryonic cerebral cortex, which may be 9-O-acetylsialylated GT3 ganglioside (Hirabayashi Y, Hirota M, Suzuki Y, Matsumoto M, Obata K, Ando S (1989) Neurosci Lett 106:193-98). In this study we established a mouse monoclonal antibody, 493D4, that recognizes 9-O-acetyl GT3 ganglioside, but not non-O-acetyl gangliosides. This antibody also reacted with 9-O-acetyl GD3 to a much lesser extent. By using this antibody, we found that O-acetyl GT3 as well as O-acetyl GD3 were expressed strongly in fetal murine cerebral cortex and decreased to an undetectable level after birth. With the assistance of TLC-immunostaining using 493D4 together with Q-Sepharose column chromatography, O-acetyl gangliosides of bovine brain were purified and the structural analysis showed the presence of O-acetyl GD3, O-acetyl LD1, O-acetyl GD2 and O-acetyl GD1b in the adult brain as extremely minor components. Interestingly, the antibody 493D4 could detect O-acetyl sialoglycoproteins in rat brain tissues. One of the major immunoreactive proteins was shown to be synaptophysin, an integral membrane protein specifically present in synaptic vesicles. This monoclonal antibody was therefore useful for sensitive detection of both O-acetylated gangliosides and glycoproteins with O-acetylated sialic acids.

Acetylation↗
PubMed1997

Elevation of [Ca2+]i of renal proximal tubular cells and down-regulation of mRNA of PTH-PTHrP, V1a and AT1 receptors in kidney of diabetic rats.

An elevation in intracellular calcium ([Ca2+]i) in rats with chronic renal failure and elevated blood levels of PTH is associated with down-regulation of the mRNA of many proteins. Similarly, in phosphate depleted animals that have normal renal function and low blood levels of PTH, [Ca2+]i is elevated and the mRNA of PTH-PTHrP receptor is down-regulated. The effect of elevation in [Ca2+]i on molecular machinery of many proteins may represent a generalized phenomenon. Diabetes mellitus may also be associated with a rise in [Ca2+]i and therefore down-regulation of the mRNA of proteins may also occur. The present study examined the effect of streptozotocin-induced diabetes mellitus in rats on the [Ca2+]i of the renal proximal tubular cells and on their mRNAs of the PTH-PTHrP, V1a and AT1 receptors. The basal levels of [Ca2+]i of these cells increased significantly (P < 0.01) after one day of diabetes and remained elevated thereafter. There was a significant (r = 0.67, P < 0.01) direct correlation between the [Ca2+]i of the cells and blood levels of glucose up to 350 mg/dl, and the value of [Ca2+]i plateaued with higher concentrations of glucose. Three days of amlodipine therapy prevented and reversed the elevated levels of [Ca2+]i despite marked hyperglycemia. The mRNA of all three receptors in the kidney were down-regulated and this defect was prevented by amlodipine which normalized the [Ca2+]i of the cells. The results show that: (1) the hyperglycemia of IDDM in rats causes a significant elevation in the basal levels of [Ca2+]i of the renal proximal tubular cells and down-regulation of their mRNA of PTH-PTHrP, V1a and AT1 receptors; (2) normalization of the [Ca2+]i of these cells by treatment of the diabetic rats with amlodipine prevented the elevation of [Ca2+]i and the down-regulation of the mRNA of these receptors; (3) these effects occurred in the presence of normal renal function and normal blood of PTH and phosphorus.

Amlodipine↗
PubMed1997

A negative regulatory region containing a glucocorticosteroid response element (nGRE) in the human interleukin-1beta gene.

Interleukin-1 beta (IL-1beta) is one of the most important inflammatory mediators in human inflammatory and immunological diseases. The regulation of human IL-1beta gene expression has been studied for several years, and a few regulatory elements have been discovered in the promoter region. However, little is known about negative regulation of IL-1beta expression at the transcriptional level, which may play an important role in anti-inflammatory and immunosuppressive effects. We have identified a negative regulatory element located in the region between -685 and -395. Within this region, a 19-bp nuclear factor binding site (-570 to -552) was characterized by DNase I footprinting and electromobility shift assay. A consensus sequence for a negative glucocorticoid response element (nGRE) and a transcription activator protein-2 binding site were noted within this footprint. Functional studies showed a 2.5-fold increase in promoter activity when this 19-bp binding site was deleted in the reporter constructs IL-1beta/CAT and IL-1beta/SV40 promoter/CAT. Dexamethasone (10(-8) M) repressed chloramphenicol acetyltransferase (CAT) production by 75% in the wild-type fragment but not in a deletion mutant lacking the 19-bp site. A protein of about 150 kD that bound to this negative regulatory sequence was identified by UV cross-linking. This is the first description of a negative regulatory region responsive to glucocorticoids in a cytokine gene.

Binding Sites↗
PubMed1997

Disruption of the clathrin heavy chain-like gene (CLTCL) associated with features of DGS/VCFS: a balanced (21;22)(p12;q11) translocation.

The smallest region of deletion overlap in the patients we have studied defines a DIGeorge syndrome/velocardiofacial syndrome (DGS/VCFS) minimal critical region (MDGCR) of approximately 250 kb within 22q11. A de novo constitutional balanced translocation has been identified within the MDGCR. The patient has some features which have been reported in individuals with DGS/VCFS, including: facial dysmorphia, mental retardation, long slender digits and genital anomalies. We have cloned the breakpoint of his translocation and shown that it interrupts the clathrin heavy chain-like gene (CLTCL) within the MDGCR. The breakpoint of the translocation partner is in a repeated region telomeric to the rDNA cluster on chromosome 21p. Therefore, it is unlikely that the patient's findings are caused by interruption of sequences on 21p. The chromosome 22 breakpoint disrupts the 3' coding region of the CLTCL gene and leads to a truncated transcript, strongly suggesting a role for this gene in the features found in this patient. Further, the patient's partial DGS/VCFS phenotype suggests that additional features of DGS/VCFS may be attributed to other genes in the MDGCR. Thus, haploinsufficiency for more than one gene in the MDGCR may be etiologic for DGS/VCFS.

Abnormalities, Multiple↗
PubMed1997

High expression of naked plasmid DNA in muscles of young rodents.

There is a time window at 2 weeks of age for achieving very high levels of foreign gene expression from the intramuscular injection of naked plasmid DNA in mice and rats. The highest expression, over 1 microg of luciferase protein/muscle, was obtained in Balb/C mice using constructs containing the CMV promoter, a chimeric intron and the luc+ luciferase gene. Approximately 50% of the myofibers were intensely blue following the intramuscular injection of a beta-galactosidase expression vector in 2 week old Balb/C mice. The effects of age, mouse strain and construct were multiplicative, resulting in >1000-fold greater luciferase and approximately 20-fold more beta-galactosidase-positive cells. These high levels of expression were unstable and were not observed in larger animals (dog, rhesus monkey). These results indicate that enormous levels of foreign gene expression can be obtained in muscle with naked DNA in vivo and will enable the temporary effects of gene function and expression in rodent muscle to be expeditiously studied.

Age Factors↗
PubMed1997

Salmonella infection increases porcine antibacterial peptide concentrations in serum.

PR-39 is a multifunctional neutrophil peptide involved in host defense and inflammation. To investigate the involvement of PR-39 in a Salmonella choleraesuis infection, a PR-39 enzyme immunoassay was developed. The concentrations of PR-39 in serum were 13.6 +/- 1.9 ng/ml before challenge and increased (P < 0.01) threefold by 10 to 14 days postinfection. Peripheral blood neutrophil counts paralleled the changes in the concentrations of PR-39 in serum, both returned to basal values by 4 weeks postinfection. These findings suggest that the concentrations of serum PR-39 reflect the involvement of this antibacterial peptide in the host's response to an S. choleraesuis infection.

Animals↗
PubMed1997

Conserved motifs II to VI of DNA helicase II from Escherichia coli are all required for biological activity.

There are seven conserved motifs (IA, IB, and II to VI) in DNA helicase II of Escherichia coli that have high homology among a large family of proteins involved in DNA metabolism. To address the functional importance of motifs II to VI, we employed site-directed mutagenesis to replace the charged amino acid residues in each motif with alanines. Cells carrying these mutant alleles exhibited higher UV and methyl methanesulfonate sensitivity, increased rates of spontaneous mutagenesis, and elevated levels of homologous recombination, indicating defects in both the excision repair and mismatch repair pathways. In addition, we also changed the highly conserved tyrosine(600) in motif VI to phenylalanine (uvrD309, Y600F). This mutant displayed a moderate increase in UV sensitivity but a decrease in spontaneous mutation rate, suggesting that DNA helicase II may have different functions in the two DNA repair pathways. Furthermore, a mutation in domain IV (uvrD307, R284A) significantly reduced the viability of some E. coli K-12 strains at 30 degrees C but not at 37 degrees C. The implications of these observations are discussed.

Adenosine Triphosphatases↗
PubMed1997

Studies of coat protein-mediated resistance to tobacco mosaic tobamovirus: correlation between assembly of mutant coat proteins and resistance.

Coat protein-mediated resistance (CP-MR) has been widely used to protect transgenic plants against virus diseases. To characterize the mechanisms of CP-MR to tobacco mosaic tobamovirus (TMV) we developed mutants of the coat protein that affected subunit-subunit interactions. Mutant CPs were expressed during TMV replication as well as in transgenic Nicotiana tabacum plants. The mutation T42-->W increased protein aggregation and T28-->W abolished aggregation and assembly, while the mutations T28-->W plus T42-->W and T89-->W altered normal CP subunit-subunit interactions. The mutant T28W was unable to assemble virus-like particles (VLPs) during infection and in transgenic plants failed to aggregate; this protein conferred no protection against challenge of transgenic plants by TMV. The mutant T42W had strong CP subunit-subunit interactions and formed VLPs but not infectious virions. Transgenic lines with this protein exhibited stronger protection against TMV infection than transgenic plants that contained the wild-type (wt) CP. It is proposed that increased resistance conferred by the T42W mutant results from strong interaction between transgenic CP subunits and challenge virus CP subunits. CP carrying the mutation T89-->W formed flexuous and unstable VLPs whereas the double mutant T28W:T42W formed open helical structures that accumulated as paracrystalline arrays. In transgenic plants, T89W and the double mutant CPs showed reduced ability to aggregate and provided lower protection against TMV infection than wt CP. A strong correlation between normal CP subunit-subunit interactions and CP-MR is observed, and a model for CP-MR involving interactions between the transgenic CP and the CP of the challenge virus as well as interference with virus movement is discussed.

Capsid↗
PubMed1997

Constitutive expression of inducible nitric oxide synthase in the mouse ileal mucosa.

It has been demonstrated previously that the inducible isoform of nitric oxide synthase (iNOS) is present throughout the intestinal tract in various inflammatory disease processes. Here we demonstrate that iNOS mRNA is present in the ileum but not in the jejunum or colon of normal mice. By Western blot analysis, iNOS protein is also detected in normal ileum, but not in the normal jejunum. However, by 3 h postinjection of 0.5 mg/kg lipopolysaccharide (LPS), iNOS mRNA is also detectable in the jejunum and colon. The enzyme message and protein, localized immunohistochemically by in situ hybridization and iNOS expression, is normally restricted to the villus epithelial cells. The iNOS mRNA was also present in the ilea of mice with defined intestinal flora (anaerobes only), germ-free mice, nude mice, and to a lesser extent in mice with severe combined immunodeficiency. These results suggest that the constitutive presence of iNOS in ileal epithelium indicates a role for this enzyme in maintaining intestinal homeostasis.

Animals↗
PubMed1997

Parathyroidectomy does not affect mRNA of PTH-PTHrP receptor in kidney, liver, and heart.

Elevated blood levels of cytosolic calcium--[Ca2+]i--appear to mediate the downregulation of the mRNA of the PTH-PTHrP receptor in states with chronic elevation of blood levels of parathyroid hormone (PTH). Data on the effect of hypoparathyroidism on basal levels of [Ca2+]i are not available, while those on the mRNA of the PTH-PTHrP receptor are variable. The present study was performed to measure [Ca2+]i in renal cells, to estimate the concentrations of mRNA of the PTH-PTHrP receptor in kidney, liver, and heart, and to examine the phosphaturic response to PTH and adenine 3',5'-cyclic monophosphate (cAMP) in normal rats as well as in animals after 3 weeks of parathyroidectomy (PTX). The basal levels of [Ca2+]i of the renal proximal tubular cells in PTX animals (163 +/- 5.1 nM) were not different from those in normal rats (160 +/- 4.9 nM). The concentrations of the mRNAs of the PTH-PTHrP receptor in kidney, liver, and heart were not upregulated after PTX. The phosphaturic responses to PTH or cAMP in normal and PTX rats were not different. The results show that PTX of rats with the consequent decrease in their blood levels of PTH is not associated with changes in the basal levels of [Ca2+]i, the concentrations of the mRNA of the PTH-PTHrP receptor, and in the phosphaturic response to PTH. These data lend support to the notion that the regulation of the mRNA of the PTH-PTHrP receptor in the presence of excess PTH or its lack is mediated through changes in basal levels of [Ca2+]i.

Animals↗
PubMed1997

Contraction kinetics of red muscle in scup: mechanism for variation in relaxation rate along the length of the fish.

We studied possible mechanisms for the twofold difference in red muscle relaxation times between the posterior (207.2 ms) and anterior (98.4 ms) musculature of scup Stenotomus chrysops, which has been shown to have a large effect on power generation during swimming. This difference was not due to contamination of the anterior bundles with faster fiber types, as histological examination showed that all bundles contained more than 98.9% red fibers. Further, maximum velocities of shortening (Vmax) at 20 degrees C were nearly identical, 5.37 MLs-1 (where ML is muscle length) for the anterior musculature and 5.47 MLs-1 for the posterior musculature, suggesting that the difference in relaxation times was not due to a difference in the crossbridge detachment rates associated with different myosin isoforms. The possibility of differences in the Ca2+ pumping rate influencing relaxation rate was explored using cyclopiazonic acid (CPA), a sarcoplasmic reticulum (SR) Ca(2+)-ATPase inhibitor. The concentration of CPA could be adjusted to slow the relaxation rate of an anterior muscle to that of a posterior muscle. However, SDS gels showed no difference in the intensity of SR Ca(2+)-ATPase protein bands between muscle positions. These results suggest that differences in the Ca2+ pumping could account for the observed difference in relaxation rate, but do not support the simplest hypothesis that the difference in relaxation rates is due to differences in numbers of Ca2+ pumps. Other possible mechanisms for this difference are explored.

Animals↗
PubMed1997

Early detection of apoptosis using a fluorescent conjugate of annexin V.

Apoptosis of mammalian cells is accompanied by various morphological changes including nuclear condensation, DNA fragmentation and cell surface changes. Methods developed over the past few years have focused on detection of DNA-associated changes that occur rather late in apoptosis. However, detection of apoptosis at early stages, before gross morphological changes, is critical for understanding the pathways of programmed cell death. In this report, we describe a rapid and reliable assay for detecting early stages of apoptosis. This assay is based on the observation that soon after initiating apoptosis, most mammalian cell types translocate phosphatidylserine (PS) from the inner face of the plasma membrane to the cell surface. Once on the cell surface, PS can be specifically detected by staining with fluorescein isothiocyanate (FITC)-labeled annexin V (annexin V-FITC), a protein with a strong, natural affinity for PS. Using this assay, we have detected apoptotic cells in culture, in real time, using fluorescence microscopy and flow cytometry. In combination with vital dye staining, the progressive stages of apoptosis were observed. PS redistribution occurs earlier than DNA-associated changes and membrane leakage. In addition, PS externalization occurs during apoptosis induced by a variety of stimuli. Therefore, the annexin V binding assay provides an excellent indicator for the early stages of apoptosis.

Annexin A5↗
PubMed1997

[Clinical and pathological analysis of liver injury resulting from rifampin and isoniazid].

OBJECTIVE: To explore the relationship between clinical manifestations and pathological changes of liver injury resulting from rifampin and isoniazid. METHOD: 194 cases treated with rifampin and isoniazid, among them liver biopsies were done once in 28 cases, twice in 17 cases, were observed. RESULTS: 21 of 59 cases with positive HBVM and 13 out of 135 cases with negative HBVM showed abnormal liver function after treatment, and the elevation of ALT was in accordance with the degree of liver pathological injury. Elevated ALT and liver pathological injury returned to normal within 6 weeks after rifampin and isoniazid were stopped or reduced in doses. No significant difference was found between chronic hepatis B and rifampin, isoniazid in causing liver pathological injury, but two different points were noted in their manifestations: 1. The cells mainly caused portal inflammation were lymphocytes in the former, while eosinophils in the latter. 2. The incidence of intrahepatic cholestasis was higher in the latter. CONCLUSION: The incidence of liver injury in HBVM positive group is higher than that in HBVM negative group because liver injury exists in the former before treatment, and the mechanisms causing liver injury may be different.

Adolescent↗
PubMed1997

[Neoplasm-inhibiting effect and sensitivity-promoting effect of indomethacin in vitro].

OBJECTIVE: To explore an anti-tumor effect of indomethacin on human colon adenocarcinoma cell line HCT116, and its mechanism. METHODS: MTT, agarose gel electrophoresis, TdT and flow cytometry were used to study cell proliferation and apoptosis. RESULTS: Indomethacin significantly inhibited the proliferation of HCT116 cells and induced apoptosis with a "dose-time" dependent pattern. A synergic effect of inhibiting cancer cells proliferation was observed when combined with 5-Fu. CONCLUSION: Inhibition of proliferation and induction of apoptosis contribute to the mechanisms for anti-tumor activity of indomethacin.

Adenocarcinoma↗