Biomedical subjects

B Zhang

Publications and source records attributed to B Zhang.

At least 415 records · Page 23Linked to original sources

PubMed1998

Experimental study on the scavenging effects of ginsenosides on oxygen free radicals using model of heterotopic heart transplantation in rats.

Oxygen-derived free radicals play an important role in myocardial injury associated ischemia and reperfusion. To investigate whether ginsenosides, as an additive agent of cardioplegic solution, can decrease toxicity of oxygen free radicals in myocardial injury, a heterotopic heart transplantation model in Wistar rats was employed. St. Thomas II cold cardioplegia containing ginsenosides 80 mg/l was used in the experimental group. St. Thomas II cold cardioplegia alone was used in the control group. After global ischemia for 60 minutes and reperfusion for 30 minutes of a transplanted heart, SOD activity in the myocardium treated with ginsenosides was significantly higher than that in the control group (N=10, p< 0.01), whereas the MDA in the myocardium treated with ginsenosides were markedly lower than that of the control group (p< 0.01). The amounts of oxygen free radicals in the myocardium treated with ginsenosides were significantly lower than that of the control group (p <0.001). This study demonstrates that ginsenosides, as a proper additive agent of cardioplegic solution, can decrease toxicity of oxygen free radicals, suggesting one of the mechanisms for its protective effects against myocardial ischemia and reperfusion injury.

Animals↗

PubMed1997

Structural determinants required for the interaction between Rho GTPase and the GTPase-activating domain of p190.

The Rho family small GTP-binding proteins are subjected to regulation by Rho GTPase-activating proteins (GAPs) in the course of transmitting diverse intracellular signals. To understand the mechanism of GAP-catalyzed GTP hydrolysis of Rho GTPases, we have studied the interaction between RhoA and p190, the RasGAP binding phosphoprotein which has been implicated as a Rho-specific GAP, by delineating the structural determinants of RhoA and p190 GAP domain (p190GD) that are involved in their functional coupling. Besides the conserved residues Tyr34, Thr37, and Phe39 in the switch I region of RhoA which are required for p190GD interaction, chimeras made between RhoA and Cdc42, a close relative of RhoA with which p190GD interacts 50-fold less efficiently, revealed that residues outside the switch I and neighboring regions of RhoA, residues 85-122 in particular, contain the major p190GD-specifying determinant(s). Mutation of the unique Asp90 of RhoA in this region mostly abolished p190GD stimulation, whereas the corresponding reverse mutation of Cdc42 (S88D) was able to respond to p190GD-catalysis similarly as RhoA. Further kinetic analysis of these mutants provided evidence that Asp90 of RhoA contributes primarily to the specific binding interaction with p190GD. On the other hand, two charged residues of p190GD, Arg1283 and Lys1321, which are located in the putative G-protein binding helix pocket of GAP domain, were found to be involved in different aspects of interaction with RhoA. The R1283L mutant of p190GD lost GAP activity but retained the ability to bind to RhoA, while K1321A failed to stimulate and to bind to RhoA. These results indicate that residue Asp90 constitutes the second GAP-interactive site in RhoA which is mostly responsible for conferring p190GD-specificity, and suggest that the role of p190GD in the GTPase reaction of RhoA is in part to supply active site residue Arg1283 for efficient catalysis.

Cell Line↗

PubMed1997

A conserved RNA-binding protein that regulates sexual fates in the C. elegans hermaphrodite germ line.

The nematode Caenorhabditis elegans has two sexes, males and hermaphrodites. Hermaphrodites Initially produce sperm but switch to producing oocytes. This switch appears to be controlled by the 3' untranslated region of fem-3 messenger RNA. We have now identified a binding factor (FBF) which is a cytoplasmic protein that binds specifically to the regulatory region of fem-3 3'UTR and mediates the sperm/oocyte switch. The RNA-binding domain of FBF consists of a stretch of eight tandem repeats and two short flanking regions. This structural element is conserved in several proteins including Drosophila Pumilio, a regulatory protein that controls pattern formation in the fly by binding to a 3'UTR. We propose that FBF and Pumilio are members of a widespread family of sequence-specific RNA-binding proteins.

Amino Acid Sequence↗

PubMed1997

Neurofilaments and orthograde transport are reduced in ventral root axons of transgenic mice that express human SOD1 with a G93A mutation.

Mice engineered to express a transgene encoding a human Cu/Zn superoxide dismutase (SOD1) with a Gly93 --> Ala (G93A) mutation found in patients who succumb to familial amyotrophic lateral sclerosis (FALS) develop a rapidly progressive and fatal motor neuron disease (MND) similar to amyotrophic lateral sclerosis (ALS). Hallmark ALS lesions such as fragmentation of the Golgi apparatus and neurofilament (NF)-rich inclusions in surviving spinal cord motor neurons as well as the selective degeneration of this population of neurons were also observed in these animals. Since the mechanism whereby mutations in SOD1 lead to MND remains enigmatic, we asked whether NF inclusions in motor neurons compromise axonal transport during the onset and progression of MND in a line of mice that contained approximately 30% fewer copies of the transgene than the original G93A (Gurney et al., 1994). The onset of MND was delayed in these mice compared to the original G93A mice, but they developed the same neuropathologic abnormalities seen in the original G93A mice, albeit at a later time point with fewer vacuoles and more NF inclusions. Quantitative Western blot analyses showed a progressive decrease in the level of NF proteins in the L5 ventral roots of G93A mice and a concomitant reduction in axon caliber with the onset of motor weakness. By approximately 200 d, both fast and slow axonal transports were impaired in the ventral roots of these mice coincidental with the appearance of NF inclusions and vacuoles in the axons and perikarya of vulnerable motor neurons. This is the first demonstration of impaired axonal transport in a mouse model of ALS, and we infer that similar impairments occur in authentic ALS. Based on the temporal correlation of these impairments with the onset of motor weakness and the appearance of NF inclusions and vacuoles in vulnerable motor neurons, the latter lesions may be the proximal cause of motor neuron dysfunction and degeneration in the G93A mice and in FALS patients with SOD1 mutations.

Age of Onset↗

PubMed1997

Regulation of experimental autoimmune encephalomyelitis by natural killer (NK) cells.

In this report, we establish a regulatory role of natural killer (NK) cells in experimental autoimmune encephalomyelitis (EAE), a prototype T helper cell type 1 (Th1)-mediated disease. Active sensitization of C57BL/6 (B6) mice with the myelin oligodendrocyte glycoprotein (MOG)35-55 peptide induces a mild form of monophasic EAE. When mice were deprived of NK cells by antibody treatment before immunization, they developed a more serious form of EAE associated with relapse. Aggravation of EAE by NK cell deletion was also seen in beta 2-microglobulin-/- (beta 2m-/-) mice, indicating that NK cells can play a regulatory role in a manner independent of CD8+ T cells or NK1.1+ T cells (NK-T cells). The disease enhancement was associated with augmentation of T cell proliferation and production of Th1 cytokines in response to MOG35-55. EAE passively induced by the MOG35-55-specific T cell line was also enhanced by NK cell deletion in B6, beta 2m-/-, and recombination activation gene 2 (RAG-2)-/- mice, indicating that the regulation by NK cells can be independent of T, B, or NK-T cells. We further showed that NK cells inhibit T cell proliferation triggered by antigen or cytokine stimulation. Taken together, we conclude that NK cells are an important regulator for EAE in both induction and effector phases.

Amino Acid Sequence↗

PubMed1997

Peptide bond formation by in vitro selected ribozymes.

An attractive solution to the problem of the origin of protein synthesis in an evolving 'RNA world' involves catalysis by nucleic acid without assistance from proteins. Indeed, even the modern ribosome has been considered to be fundamentally an RNA machine, and the large ribosomal subunit can carry out peptidyl transfer in the absence of most of its protein subunits. Successive cycles of in vitro selection and amplification have been used to find RNAs that perform many biochemical reactions, including transfer of an RNA-linked amino acid to their own 5'-amino-modified terminus. Here we demonstrate the in vitro selection of ribozymes (196 nucleotides) that perform the same peptidyl transferase reaction as the ribosome: that is, they can join amino acids by a peptide bond. Like ribosome substrates, one amino acid (N-blocked methionine) is esterified to the 3'(2')-O of adenosine, whereas the acceptor amino acid (phenylalanine) has a free amino group. Our best characterized ribozyme recognizes the amino-acid ester substrate by binding its adenosine moiety, and is therefore capable of utilizing Leu- and Phe- as well as Met-derived substrates. Such lack of specificity with respect to the amino acid is a feature necessary for a generalized protein-synthesizing enzyme.

Adenosine Monophosphate↗

PubMed1997

Chromosomal aberrations and micronuclei in lymphocytes of workers at a phosphate fertilizer factory.

The frequencies of chromosomal aberrations (CA) and micronuclei (MN) in peripheral blood lymphocytes of 40 workers at a phosphate fertilizer factory in North China, were studied. HF and SiF4 are the main air pollutants and small amounts of dust containing fluoride, NH3 and SO2 were also present in the factory. It was shown that the chemicals caused an increase in both CA and MN. The mean frequencies per 100 metaphase of major CA type (chromosome rings, translocations, and dicentrics) of the workers and the non-exposed controls were 0.91 and 0.24 (p < 0.01), respectively. The average percentages of lymphocytes with MN of the workers and the controls were 1.55 +/- 0.71 and 0.62 +/- 0.54 (p < 0.01), respectively. Both CA frequency and MN frequency of the workers increased with length of the chemical exposure period up to 10 years.

Air Pollutants, Occupational↗

PubMed1997

Storing analog data in a video record.

We have designed a simple device that will encode, in machine readable form, multiple analog data in a video record. The analog data is visible to the user within the video frame permitting visual correlation of these signals with activity observed in the image. By superimposition of both analog and video data sets, the two are tightly synchronized and remain so in all copies of the data. The analog data can be separated from the image following digitization by a frame grabber. The bandwidth for each of the five analog channels is approximately 5 kHz. The device, which is essentially an eight channel video multiplexor, includes a video channel, a field counter, an amplitude calibration signal and five analog data channels. The amplitude calibrator allows corrections for gain errors that are particularly prevalent when data is stored on video tape.

Calibration↗

PubMed1997

Autoantibodies to IA-2 and IA-2 beta in insulin-dependent diabetes mellitus recognize conformational epitopes: location of the 37- and 40-kDa fragments determined.

IA-2 and IA-2 beta are major autoantigens in insulin-dependent diabetes mellitus (IDDM) and the precursors, respectively, of a 40-and 37-kDa tryptic fragment that reacts with IDDM sera. In the present study, by amino acid sequencing of recombinant IA-2 and IA-2 beta, we determined the tryptic cleavage sites involved in the generation of these fragments. Both cleavage sites are immediately after an arginine residue at position 653 for IA-2 and position 679 for IA-2 beta. The resulting tryptic fragments are 326 and 307 amino acids in length and retain their ability to react with IDDM sera. In contrast to IA-2 and IA-2 beta, other members of the protein tyrosine phosphatase (PTP) family (i.e., RPTP kappa, RPTPmu, NU-3, SHP, and 3CH134) are completely susceptible to digestion by trypsin. Sequence analysis revealed five conserved cysteine residues in IA-2 and IA-2 beta that are not present in other PTPs. Reduction and alkylation of IA-2 and IA-2 beta recombinant proteins resulted in loss of both resistance to digestion by trypsin and reactivity with autoantibodies in IDDM sera. It is concluded that disulfide bond formation plays a critical role in the maintenance of antigenic structure and that the autoantibodies to IA-2/IA-2 beta in IDDM sera recognize conformational epitopes.

Amino Acid Sequence↗

PubMed1997

Characterization of the interactions between the small GTPase Cdc42 and its GTPase-activating proteins and putative effectors. Comparison of kinetic properties of Cdc42 binding to the Cdc42-interactive domains.

The small GTPase Cdc42 interacts with multiple factors to transduce diverse intracellular signals. The factors that preferentially recognize the GTP-bound, active state of Cdc42 include a panel of GTPase-activating proteins (GAPs), the Cdc42/Rac interactive binding (CRIB) motif-containing molecules, and the RasGAP domain containing IQGAP1 and IQGAP2. In the present study, we have determined the kinetic parameters underlying the functional interactions between the Cdc42-binding domains of some of these factors and Cdc42 by monitoring the continuous release of gammaPi and have compared the ability of the domains to bind to Cdc42. The catalytic efficiencies (Kcat/Km) of the GAP domains of Bcr, 3BP-1, and p190 on Cdc42 are found to be 60-, 160-, and over 500-fold less than that of Cdc42GAP, respectively, and the differences are due, to a large part, to differences in Km. The Km values of the GAP domains compare well to the binding affinity to the guanylyl imidodiphosphate-bound Cdc42, suggesting a rapid equilibrium reaction mechanism. The affinity of the Cdc42-binding domains of the CRIB motif of Wiskott-Aldrich Syndrome protein and p21(cdc42/rac)-activated kinase 1, and the RasGAP-related domain of IQGAP1, which all inhibit the intrinsic rate of GTP hydrolysis of Cdc42, are found to be 4, 0.7, and 0.08 microM, respectively. These quantitative analysis provide insight that Cdc42GAP functions as an effective negative regulator of Cdc42 by fast, relatively tight binding to the GTP-bound Cdc42, whereas IQGAP1 interacts with Cdc42 as a putative effector with over 10-fold higher affinity than the CRIB domains and GAPs, and suggest that various GAPs and effectors employ distinct mechanism to play roles in Cdc42-mediated signaling pathways.

Binding Sites↗

PubMed1997

Association of DNA with nuclear matrix in in vitro assembled nuclei induced by rDNA from Tetrahymena shanghaiensis in Xenopus egg extracts.

Nuclei assembled in vitro from Xenopus egg extracts and DNA display many properties known from in vivo nuclei. However, the distribution pattern of DNA in such nuclei remains unknown. We introduced rDNA from Tetrahymena macronuclei into Xenopus egg extracts and examined the association of specific DNA sequences with the nuclear matrix (NM) of the nuclei formed. We found that the 5' non-transcribed spacers (5'-NTS) specifically bound to the NM as previously shown in normal Tetrahymena macronuclei.

Animals↗

PubMed1997

Ischaemia-reperfusion injury in mouse skeletal muscle is reduced by N omega-nitro-L-arginine methyl ester and dexamethasone.

We have developed a model of ischaemia-reperfusion injury in C57BL/6 mice involving ischaemia for 0.5 to 2.5 h with an elastic tourniquet on one hind limb and reperfusion for 24 h, analogous to a well-established model of ischaemia-reperfusion injury in the rat. Viability was assessed in tissue homogenates of the gastrocnemius muscles from the affected and contralateral control limb by a triphenyl tetrazolium chloride dye reaction, measuring the activity of the oxidative mitochondrial enzymes. After 1.5 h ischaemia and 24 h reperfusion, viability in the ischaemic-reperfused limb was 13%, with the control muscle regarded as 100% viable. Significant improvements in viability to 86% (P < 0.05) and 56% (P < 0.05) were achieved, with administration 30 min prior to tourniquet release, of the nitric oxide (NO) synthase inhibitor nitro-L-arginine methyl ester (L-NAME, 30 mg/kg) and the anti-inflammatory glucocorticoid dexamethasone (2.5 mg/kg) respectively, with similar findings in the rat tourniquet model.

Animals↗

PubMed1997

Diagnostic value of immunochemical fecal occult blood test for small colorectal neoplasms.

This study was designed to assess the diagnostic value of an immunochemical fecal occult blood test for small colorectal adenomas. In a screening program based comparative study, 60 colorectal adenomas under 1 cm (small polyp group), 60 colorectal adenomas 1 cm or larger (large polyp group) and 60 colorectal cancers (cancer group) detected by occult blood screening, and 120 healthy controls (control group), served as subjects. Moreover, 50 small adenomas, 50 large adenomas and 50 cancers detected in out-patient clinics, and 100 controls, served as subjects of a hospital based comparative study. Each of these subjects was tested by an immunochemical occult blood with 3 consecutive days, after removal of these colorectal neoplasms in a screening program based study and before as well as after removal in a hospital based study, and the positivity rate of this test was evaluated among these four groups in the two comparative studies. The positivity rate in a screening program based study was 23% for small polyp group, 8% for large polyp group, 5% for cancer group and 6% for control group, respectively. Significant difference was noted between small polyp and cancer as well as control groups (p <0.01), and between small polyp and large polyp groups (p <0.05). In a hospital based study, however, there was no significant difference in the positivity rate among four groups. These results suggest that high proportion of small polyps detected by fecal occult blood test screening may not be associated with bleeding from adenomas but probably from another gastrointestinal pathology.

Adenoma↗

PubMed1997

Interaction of sodium chloroplatinate and iproplatin with metallothionein in vivo.

The abilities of platinum(IV) complexes to induce the biosynthesis of metallothionein (MT) were investigated in rabbits given injections s.c. of sodium chloroplatinate (Na2PtCl6) and iproplatin (cis-dichloro-bis-isopropylamine-trans-dihydroxylplatinum IV). It is revealed for the first time that both complexes can induce MT synthesis in the liver and the kidney, but the induction ability was weaker compared to Zn2+ compounds. The induced MT was purified and identified. The hepatic MT resulting from Na2PtCl6 injection only contained Zn, whereas the hepatic MT from iproplatin injection and the renal MT from injection of both complexes contained 4-5 Zn and 1-2 Pt per mole of protein, and the renal MT also contained 1-2 Cu per mole of protein. The oxidation state of platinum in the MT is +2 as determined by X-ray photoelectron spectroscopic measurements. Pretreatment with Zn(NO3)2 elevated the levels of MT, but the binding of Pt to MT was significantly less compared to that without Zn(NO3)2 pretreatment. The data obtained from the amino acid composition analysis were consistent with the theoretical values. Upon these bases, the role of MT in relation to its involvement in the metabolism of Pt(IV) complexes and the mechanism of drug resistance to the Pt(IV) complexes as antitumor agents are discussed.

Animals↗

PubMed1997

Reaction of a platinum(IV) complex with native Cd,Zn-metallothionein in vitro.

The first observation of a redox process following a substitution reaction between a platinum(IV) complex K2PtCl6 with rabbit liver native Cd,Zn-MT is presented. The reaction features and products are studied by UV-visible and circular dichroism spectroscopy, chromatography, and X-ray photoelectron spectroscopic measurements. It is a significant complicated reaction comprising redox and substitution reactions. The reaction generates monomeric and dimeric products, and higher oligomers precipitate with intra- or intra- and intermolecular CyS-SCy linkages. Pt(IV) is reduced to Pt(II), which then binds to the monomeric and dimeric products, and may also bind to higher oligomers. The beta-cluster is more reactive than the alpha-cluster, and reacts first with K2PtCl6. Cd5Pt2 and Cd4Pt4 were found when native Cd,Zn-MT reacted with 2 and 4 molar equivalents of Pt(IV) for 2 h in which four Cd ions were located in the alpha-cluster. The amounts of Cd and Pt ions decreased in both monomeric and dimeric products when the reaction was prolonged and intramolecular CyS-SCy linkages increased. Besides the oligomers which precipitated, only dimeric products were formed when the reaction molar ratio of Pt(IV) to MT was more than 10:1. Cd3Pt6 and Cd1Pt8 were obtained when the reaction occurred for 2 and 72 h, respectively. The structure of the clusters may exist when native Cd, Zn-MT reacts with substoichiometric quantities of K2PtCl6 (< 0.5 K2PtCl6 per MT thiolate) for a short time (2 h), but may be partly disrupted with stoichiometric or excess quantities of K2PtCl6 (> or = 0.5 K2PtCl6 per MT thiolate) for a long time. The disruption of the cluster structures results in an increase of the nonbridge thiolate and an increase of the binding sites to Pt ions. The mechanism of the antitumor activity and developing drug resistance of Pt(IV) complex drugs is discussed.

Animals↗

PubMed1997

Inhibition of sickle erythrocyte adhesion to immobilized thrombospondin by von Willebrand factor under dynamic flow conditions.

Sickle red blood cell (RBC) adhesion to the blood vessel wall is hypothesized to be the initiating event in the periodic vaso-occlusive episodes that characterize sickle cell disease (SCD). Thrombospondin-1 (TSP) and von Willebrand factor (vWF) have each been implicated in the adhesion of sickle RBC to vascular endothelial cells (EC) and subendothelial matrices. To better understand the contributions of each of these adhesive glycoproteins, we examined the adhesion of sickle RBC to immobilized TSP and vWF using a parallel plate flow chamber. Under postcapillary venular shear stress (1 dyne/cm2), sickle RBC adhered preferentially to TSP. To explore potential interactive effects of vWF and TSP, we examined sickle RBC adhesion to mixtures of these proteins. Whether the proteins were first combined in solution or sequentially applied to the slide, the presence of vWF inhibited the binding of sickle RBC to TSP. The inhibition of adhesion by vWF was shown to be the result of specific and saturable binding of vWF to TSP. Furthermore, vWF in solution at normal plasma levels also inhibited RBC adhesion to immobilized TSP. These data indicate that sickle RBC adhesion in vivo may be significantly influenced by the relative concentrations of TSP and vWF in the vascular wall.

Anemia, Sickle Cell↗

PubMed1997

Functional antagonism between CCAAT/Enhancer binding protein-alpha and peroxisome proliferator-activated receptor-gamma on the leptin promoter.

The ob gene product, leptin, is a major hormonal regulator of appetite and fat cell mass. Recent work has suggested that the antidiabetic agents, the thiazolidinediones (TZ), which are also high affinity ligands of peroxisome proliferator-activated receptor-gamma (PPARgamma), inhibit leptin expression in rodents. To examine the effects of this class of drug on the leptin gene in adipocytes we performed Northern analysis on primary rat adipocytes cultured in the presence or absence of TZ. TZ reduced leptin mRNA levels by 75%. To determine whether this effect was mediated at the transcriptional level, we isolated 6510 base pairs of 5'-flanking sequence of the leptin promoter and studied reporter constructs in primary rat adipocytes and CV-1 cells. Sequence analysis demonstrated the presence of a consensus direct repeat with a 1-base-pair gap site between -3951 and -3939 as well as a consensus CCAAT/enhancer binding protein (C/EBP) site between -55 and -47. Our functional analysis in transfected primary rat adipocytes demonstrates that, despite the presence of a canonical direct repeat with a 1-base-pair gap site, TZ alone decreases reporter gene expression of leptin promoter constructs ranging from -6510 to +9 to -65 to +9. In CV-1 cells, which contain endogenous PPARgamma, TZ treatment alone had little effect on these constructs. However, TZ treatment did inhibit C/EBPalpha-mediated transactivation of the leptin promoter. This down-regulation of leptin reporter constructs mapped to a -65 to +9 promoter fragment which binds C/EBPalpha in gel-mobility shift assays but does not bind PPARgamma2 alone or as a heterodimer with 9-cis-retinoic acid receptor. Conversely, the promoter (-5400 to +24 base pairs) of the aP2 gene, another adipocyte-specific gene, was induced 7.3-fold by TZ. Co-transfection with C/EBPalpha minimally stimulated the aP2 promoter from basal levels but notably blocked activation by TZ. These data indicate that PPARgamma and C/EBPalpha can functionally antagonize each other on at least two separate promoters and that this mechanism may explain the down-regulation of leptin expression by thiazolidinediones.

Adipocytes↗

PubMed1997

Construction and screening of a cosmid library generated from a somatic cell hybrid bearing human chromosome 15.

A cosmid library has been constructed with DNA isolated from a mouse/human hybrid cell line designated A15, which was previously characterized and shown to retain chromosome 15 as the only human material. The library was generated and stored as 34 independent pools of primary colonies at 8-10,000 colonies per pool. Screening colonies representing pools of this library by hybridization with a human-specific repetitive probe has facilitated the identification of random clones bearing human inserts. To data, 43 unique clones have been isolated and the inserts mapped by fluorescence in situ hybridization (FISH) to chromosome 15. An STS was generated for each of these clones by end sequencing of the inserts. Two of these clones, c36 and MR23, were mapped by FISH to 15q26.1, and end-sequence data revealed homology to different regions of the FES protooncogene. Sequence generated from the other end of the c36 insert was found to match the previously identified and unmapped coding sequence EST00075. In addition to the identification of such random clones, establishment of the library as pools was expected to facilitate the PCR-based identification of unique clones representing specific regions of chromosome 15. Screening of the library pools with primers specific to the FES region led to the recovery of five independent clones facilitating the development of a cosmid contig encompassing the FES gene. One of the cosmids isolated by PCR-based analysis was derived from the same pool as M23 and was subsequently shown to be identical to M23. The data offer the first report of a chromosome 15 cosmid library and demonstrate the value of utilizing pools to evaluate libraries generated from complex sources like somatic cell hybrids.

Animals↗