Biomedical subjects

C Wang

Publications and source records attributed to C Wang.

At least 361 records · Page 20Linked to original sources

PubMed2000

Atomic force microscopy analysis of intermediates in cobalt hexammine-induced DNA condensation.

The packaging pathway of cobalt hexammine-induced DNA condensation on the surface of mica was examined by varying the concentration of Co(NH3)6(3+) in a dilute DNA solution and visualizing the condensates by atomic force microscopy (AFM). Images reveal that cobalt hexammine-induced DNA condensation on mica involves well-defined structures. At 30 microM Co(NH3)6(3+), prolate ellipsoid condensates composed of relatively shorter rods with linkages between them are formed. At 80 microM Co(NH3)6(3+), the condensed features include toroids with average diameter of approximately 240 nm as well as U-shaped and rod-like condensates with nodular appearances. The results imply that the condensates, whether toroids, U-shaped or rod-like structures have similar intermediate state which includes relatively shorter rod-like segments. The average size of the condensed toroids after incubated at room temperature for 5 h (approximately 240 nm) is much larger than that incubated for 0.5 h (approximately 100 nm). The results indicate that the condensation of DNA by Co(NH3)6(3+) is a kinetic-controlled process.

Adsorption↗

PubMed2000

Lack of collagen type specificity for lysyl hydroxylase isoforms.

Lysyl hydroxylase is the enzyme catalyzing the formation of hydroxylysyl residues in collagens. Large differences in the extent of hydroxylysyl residues are found among collagen types. Three lysyl hydroxylase isoenzymes (LH1, LH2, LH3) have recently been characterized from human and mouse tissues. Nothing is known about the distribution of these isoforms within cells or whether they exhibit collagen type specificity. We measured mRNA levels of the three isoforms, as well as the mRNAs of the main collagen types I, III, IV, and V and the alpha subunit of prolyl 4-hydroxylase, another enzyme involved in collagen biosynthesis, in different human cell lines. Large variations were found in mRNA expression of LH1 and LH2 but not LH3. Immunoblotting was utilized to confirm the results of Northern hybridization. The levels of mRNA of LH1, LH2, and the alpha subunit of prolyl 4-hydroxylase showed significant correlations with each other. The LH3 mRNA levels did not correlate with those of LH1, LH2, or the alpa subunit of prolyl 4-hydroxylase, clearly indicating a difference in the regulation of LH3. No correlation was observed between LH isoforms and individual collagen types, indicating a lack of collagen type specificity for lysyl hydroxylase isoforms. Our observations suggest that LH1, LH2, and the alpha subunit of prolyl 4-hydroxylase are coregulated together with total collagen synthesis but not with the specific collagen types and indicate that LH3 behaves differently from LH1 and LH2, implying a difference in their substrates. These observations set the basis for further studies to define the functions of lysyl hydroxylase isoforms.

Animals↗

PubMed2000

Conserved composition of mammalian box H/ACA and box C/D small nucleolar ribonucleoprotein particles and their interaction with the common factor Nopp140.

Small nucleolar ribonucleoprotein particles (snoRNPs) mainly catalyze the modification of rRNA. The two major classes of snoRNPs, box H/ACA and box C/D, function in the pseudouridylation and 2'-O-methylation, respectively, of specific nucleotides. The emerging view based on studies in yeast is that each class of snoRNPs is composed of a unique set of proteins. Here we present a characterization of mammalian snoRNPs. We show that the previously characterized NAP57 is specific for box H/ACA snoRNPs, whereas the newly identified NAP65, the rat homologue of yeast Nop5/58p, is a component of the box C/D class. Using coimmunoprecipitation experiments, we show that the nucleolar and coiled-body protein Nopp140 interacts with both classes of snoRNPs. This interaction is corroborated in vivo by the exclusive depletion of snoRNP proteins from nucleoli in cells transfected with a dominant negative Nopp140 construct. Interestingly, RNA polymerase I transcription is arrested in nucleoli depleted of snoRNPs, raising the possibility of a feedback mechanism between rRNA modification and transcription. Moreover, the Nopp140-snoRNP interaction appears to be conserved in yeast, because depletion of Srp40p, the yeast Nopp140 homologue, in a conditional lethal strain induces the loss of box H/ACA small nucleolar RNAs. We propose that Nopp140 functions as a chaperone of snoRNPs in yeast and vertebrate cells.

Amino Acid Sequence↗

PubMed2000

Initiation of nucleolar assembly is independent of RNA polymerase I transcription.

This report examines the distribution of an RNA polymerase I transcription factor (upstream binding factor; UBF), pre-rRNA processing factors (nucleolin and fibrillarin), and pre-rRNAs throughout mitosis and postmitotic nucleologenesis in HeLa cells. The results demonstrate that nucleolin, fibrillarin, and pre-rRNAs synthesized at G2/M phase of the previous cell cycle are directly recruited to UBF-associated nucleolar organizer regions (NORs) early in telophase before chromosome decondensation. Unlike the fusion of prenucleolar bodies to the nucleoli, this early recruitment of processing factors and pre-rRNAs is independent of RNA polymerase I transcription. In the absence of polymerase I transcription, the initial localization of nucleolin, fibrillarin, and pre-rRNAs to UBF-associated NORs generates segregated mininucleoli that are similar to the larger ones observed in interphase cells grown under the same conditions. Pre-rRNAs are juxtaposed to UBF-nucleolin-fibrillarin caps that may represent the segregated nucleoli observed by electron microscopy. These findings lead to a revised model of nucleologenesis. We propose that nucleolar formation at the end of mitosis results from direct recruitment of processing factors and pre-rRNAs to UBF-associated NORs before or at the onset of rDNA transcription. This is followed by fusion of prepackaged prenucleolar bodies into the nucleolus. Pre-ribosomal ribonucleoproteins synthesized in the previous cell cycle may contribute to postmitotic nucleologenesis.

Cell Cycle↗

PubMed2000

Modulatory influence of feedback projections from area 21a on neuronal activities in striate cortex of the cat.

We have examined the influence of 'feedback' projections from extrastriate visual cortical area 21a on the responses of neurons in area 17 of the cat, by cooling area 21a to 5-10 degrees C while the temperature over the recording sites was kept at 36 degrees C. Orientation, direction and length selectivities as well as contrast sensitivity were tested before and during cooling and after rewarming of area 21a. Overall, for the sample of cells recorded from the part of area 17 visuotopically corresponding to area 21a, the 'spontaneous' activity (at background illumination of 1 cd/m(2)) and the responsiveness to visual stimuli of standard contrast (15) were significantly reduced by inactivation of area 21a. In about half of the cells inactivation of area 21a affected substantially the sharpness of orientation-tuning. However, only in a minority of the cells were the direction or length selectivities significantly affected by inactivation of area 21a. Thus (i) the feedback projections from area 21a appear to exert mainly an excitatory influence on the background activity and responsiveness of area 17 cells and (ii) only in subgroups of area 17 cells does the feedback activity originating from area 21a appear to modulate specifically certain receptive field properties.

Animals↗

PubMed2000

Vagally mediated sympathoexcitation and central depression by desflurane in rabbits.

The effects of desflurane on renal sympathetic nerve activity (RSNA) were studied in intact or vagotomized anaesthetized rabbits with initial concentrations of 4.5-18%, subsequently equilibrated to end-tidal concentrations from 3%, 6%, 9% and 12% each for 20 min allowing sympathetic activity to stabilize. In intact animals, immediate transient increases in mean sympathetic activity from 27% to 63% were closely related to initial concentrations from 4.5% to 18%. During subsequent equilibration this remained elevated by 25-30% up to 6%, returned to control at 9% and fell by 33% at 12%. Bilateral vagotomy abolished sympathoexcitation apart from small increases in sympathetic activity, for example 14% at 4.5% (P < 0.05). We conclude that increases in inspired desflurane concentrations evoked rapid transient vagally mediated reflex sympathoexcitation with a small extra-vagal contribution. Central depression of sympathetic activity started at 6% and was 33% below baseline at 12%.

Anesthetics, Inhalation↗

PubMed2000

Reduction of mortality in specific-pathogen-free layer chickens by a caprine serum fraction after infection with Pasteurella multocida.

Caprine serum was fractionated by size, and its proteinaceous material <8,000 Da [caprine serum fraction immunomodulator 2 (CSF-I2)] was evaluated for its ability to impart immunoresistance to specific-pathogen-free (SPF) layer chickens. The SPF layers were challenged with 18 to 30 cfu of Pasteurella multocida X-73 (serotype 1) at 5 wk of age. A high degree of mortality was apparent 24 and 48 h later (62+/-14% and 88+/-7%, respectively). Mortality observed after 48 h was minimal. Noting the rapid onset of mortality, we administered CSF-I2 (material that expressed no direct antimicrobial activity but was believed to be an immunostimulant) 1 d before challenge and coincident to time of challenge. The group of birds that received CSF-I2 (either 5 or 10 mg per administration) expressed significant reduction in mortality throughout the 1-wk study period. Reduction in mortality appeared to be dose dependent. Birds that received two administrations of 10 mg CSF-I2 had significantly fewer deaths than did the group of birds that received half that amount. No deaths were recorded through 24 h, whereas, at 48 h, the percentage mortality was 13 in CSF-I2-treated birds. This study demonstrates that one or more small molecular weight compounds isolated from caprine serum were able to reduce mortality in SPF layers infected with Pasteurella multocida.

Adjuvants, Immunologic↗

PubMed2000

Mu-class GSTs are responsible for aflatoxin B(1)-8, 9-epoxide-conjugating activity in the nonhuman primate macaca fascicularis liver.

Mice are resistant to the carcinogenic effects of the mycotoxin aflatoxin B(1) (AFB(1)) because they constitutively express an alpha-class glutathione S-transferase (mGSTA3-3) that has high (approximately 200,000 pmol/min/mg) activity toward aflatoxin B(1)-8, 9-epoxide (AFBO). Rats do not constitutively express a GST with high AFBO-conjugating activity and are sensitive to AFB(1)-induced hepatocarcinogenesis. Constitutively expressed human hepatic alpha-class GSTs (hGSTA1-1 and hGSTA2-2) possess little or no AFBO-detoxifying activity (<2 pmol/min/mg). Recently, we found that the nonhuman primate, Macaca fascicularis (Mf), exhibits significant (approximately 300 pmol/min/mg) constitutive hepatic GST activity towards AFBO. To determine which specific GST isoenzyme(s) is (are) responsible for this activity, MF: GSTs were purified from liver tissue and characterized and, Mf mu-class GST cDNAs were cloned by reverse transcriptase-coupled polymerase chain reaction (RT-PCR). Purification by glutathione agarose (GSHA) affinity chromatography yielded a protein, GSHA-GST, that exhibited relatively high AFBO-conjugating activity (239 pmol/min/mg) compared to other GST-containing peaks. Western blotting and enzymatic activity analyses revealed that GSHA-GST belongs to the mu class. Two distinct mu-class GST cDNAs, mfaGSTM1 (GenBank accession # AF200709) and mfaGSTM2 (GenBank accession # AF200710), were generated by RT-PCR. CDNA-derived amino acid sequence analysis revealed that mfaGSTM1 and mfaGSTM2 share 97% and 96% homology with the human mu-class GSTs hGSTM4 and hGSTM2, respectively. In contrast to recombinant mfaGSTM1-1, which had no detectable AFBO-conjugating activity, mfaGSTM2-2 exhibited this activity at 333 pmol/min/mg. Activity profiles for the stereoisomers exo- and endo-AFBO, and of 1-chloro-2,4-dinitrobenzene of the purified protein GSHA-GST and recombinant mfaGSTM2-2, suggested that they are two distinct enzymes. Our results indicate that, in contrast to rodents, mu-class GSTs are responsible for the majority of AFBO-conjugating activity in the liver of Macaca fascicularis.

Aflatoxin B1↗

PubMed2000

Redistribution of Bax is an early step in an apoptotic pathway leading to germ cell death in rats, triggered by mild testicular hyperthermia.

Programmed cell death occurs spontaneously during spermatogenesis and can be induced in a cell- and stage-specific manner by mild testicular hyperthermia. Studies using transgenic mice suggest the involvement of Bcl-2 proteins in regulating germ cell apoptosis. To delineate further the pathways involved, we examined the temporal changes in proapoptotic Bax and antiapoptotic Bcl-2 in rat testes after transient exposure to heat (43 degrees C for 15 min). Germ cell apoptosis, involving exclusively early (I-IV) and late (XII-XIV) stages, was activated within 6 h. Initiation of apoptosis was preceded by a redistribution of Bax from a cytoplasmic to perinuclear localization within 0.5 h of heating as assessed by immunocytochemical methods. In contrast, Bcl-2 is distributed both in the cytoplasm and nucleus in those cell types susceptible to heat-induced apoptosis. Despite the striking redistribution, Bax levels remained unchanged as determined by Western analysis; Bcl-2 levels increased significantly by 6 h after heat exposure. Reverse transcription-polymerase chain reaction analysis indicated no change in either Bax or Bcl-2 mRNA levels in response to heat, suggesting the involvement of post-transcriptional rather than transcriptional mechanisms mediating their activity. The marked subcellular redistribution of Bax prior to activation of apoptosis and the increase in Bcl-2 suggest an involvement of Bcl-2 family members in heat-induced apoptotic death of germ cells.

Animals↗

PubMed2000

Hsp72 induction: a potential molecular mediator of the delay phenomenon.

The molecular basis of enhanced ischemic tissue survival in flaps preconditioned by surgical delay is poorly understood. Because elevated expression of so-called heat shock or stress proteins has been shown to protect tissues/organs against ischemic injury, the authors examined whether the levels of the most highly induced stress protein-hsp72-were elevated in delayed muscle flaps using a rat muscle flap model. Bilateral latissimus dorsi muscle flaps based on the thoracodorsal vessels were elevated in 16 male Sprague-Dawley rats. For each animal, one side was selected randomly to undergo preconditioning by surgical delay for a 7-day period prior to elevation. Delay was accomplished by preserving the thoracodorsal pedicle and a single large distal intercostal perforating vessel. After bilateral flap elevation, latissimus dorsi tissue was harvested from proximal, central, and distal flap segments 0, 1, 3, and 7 days postoperatively (N = 4 for each group), and was analyzed for the expression of hsp72 via Western blot analysis. At the time of harvest, flap viability was assessed by staining with nitroblue tetrazolium. Flap perfusion was measured prior to muscle elevation and harvest using laser Doppler flowmetry. The results demonstrate that delayed muscle flaps had significantly greater total perfusion (p < 0.05) and survival (p < 0.03) 1, 3, and 7 days after elevation compared with the acutely elevated control tissue. Western blot analysis revealed that tissues harvested from the delayed flaps expressed substantially higher levels of hsp72 compared with the acutely elevated control samples. Segmental analysis also revealed a proximal > middle > distal expression of hsp72 in the delayed flaps (p < 0.05). Flap preconditioning by surgical delay increases the expression of hsp72. Moreover, regional differences in hsp72 gene expression are associated with differences in perfusion and survival of delayed muscle flaps. These results indicate that hsp72 may play a substantial role in mediating the delay phenomenon.

Analysis of Variance↗

PubMed2000

Effect of genistein on cardiovascular responses to angiotensin II in conscious unrestrained rats.

Soybeans contain genistein, a phytoestrogen that may have beneficial effects in coronary artery disease, osteoporosis, and breast cancer. In vitro studies demonstrated that genistein reduced vascular smooth muscle contractions to angiotensin II. We tested the hypothesis that genistein attenuates the overall cardiovascular responses to angiotensin II via nongenomic mechanisms in conscious rats. Mean arterial blood pressure (MAP) was recorded from conscious unrestrained Sprague-Dawley rats (n = 26) approximately 48 hours after surgery. Cumulative dose response curves to angiotensin II (10-200 ng/kg/min) were constructed before and after i.v. treatment with genistein given as a single bolus dose of 160 microg/kg or 1500 microg/kg, or as a loading dose of 160 microg/kg followed by an infusion at a rate of 20 microg/kg. Angiotensin II infusions were associated with graded increases in arterial pressure ranging between 0+/-1 and 35+/-4 mm Hg. These pressor responses were accompanied by significant dose-dependent decreases in heart rate. None of the genistein treatment regimens significantly affected the pressor responses to angiotensin II. Accordingly, we conclude that short-term i.v. treatment with genistein does not depress pressor responsiveness to angiotensin II.

Angiotensin II↗

PubMed2000

Laser-driven electron cyclotron autoresonance accelerator with production of an optically chopped electron beam.

Analysis is presented of the gyroresonant acceleration of electrons in a vacuum using a focused laser. Continuous and equal acceleration is shown for electrons injected at all optical phases over an interaction length of tens of centimeters. Beam stalling is avoided as beam energy increases. Acceleration from 50 to 178 MeV is predicted for a 4 TW, 10.6-microm laser focused to a waist radius of 1.0 mm; these parameters correspond to a planned experiment. A beam stop with an off-axis hole after acceleration is shown to create a train of optically chopped bunches with 3-fs bunch lengths and a 35-fs period.

Journal Article↗

PubMed2000

Involvement of phospholipase D in wound-induced accumulation of jasmonic acid in arabidopsis.

Multiple forms of phospholipase D (PLD) were activated in response to wounding, and the expressions of PLDalpha, PLDbeta, and PLDgamma differed in wounded Arabidopsis leaves. Antisense abrogation of the common plant PLD, PLDalpha, decreased the wound induction of phosphatidic acid, jasmonic acid (JA), and a JA-regulated gene for vegetative storage protein. Examination of the genes involved in the initial steps of oxylipin synthesis revealed that abrogation of the PLDalpha attenuated the wound-induced expression of lipoxygenase 2 (LOX2) but had no effect on allene oxide synthase (AOS) or hydroperoxide lyase in wounded leaves. The systemic induction of LOX2, AOS, and vegetative storage protein was lower in the PLDalpha-suppressed plants than in wild-type plants, with AOS exhibiting a distinct pattern. These results indicate that activation of PLD mediates wound induction of JA and that LOX2 is probably a downstream target through which PLD promotes the production of JA.

Arabidopsis↗

PubMed2000

Structure of human erythrocyte catalase.

Catalase (E.C. 1.11.1.6) was purified from human erythrocytes and crystallized in three different forms: orthorhombic, hexagonal and tetragonal. The structure of the orthorhombic crystal form of human erythrocyte catalase (HEC), with space group P2(1)2(1)2(1) and unit-cell parameters a = 84.9, b = 141.7, c = 232.5 A, was determined and refined with 2.75 A resolution data. Non-crystallographic symmetry restraints were employed and the resulting R value and R(free) were 0.206 and 0.272, respectively. The overall structure and arrangement of HEC molecules in the orthorhombic unit cell were very similar to those of bovine liver catalase (BLC). However, no NADPH was observed in the HEC crystal and a water was bound to the active-site residue His75. Conserved lattice interactions suggested a common growth mechanism for the orthorhombic crystals of HEC and BLC.

Animals↗

PubMed2000

ITS regions in diploids of Aegilops (Poaceae) and their phylogenetic implications.

The nucleotide sequences of the internal transcribed spacer (ITS) of nuclear ribosomal DNA in nine diploid species representing six sections of Aegilops were determined by direct sequencing of PCR-amplified DNA fragments. These sequences were aligned with two ITS sequences of additional species from Genbank. Sequence divergences were estimated using Kimura two-parameter model, and the phylogenetic analyses were performed using the maximum parsimony (MP) and the neighbor-joining (NJ) methods with PAUP and PHYLIP, respectively. The sequence divergences between the diploid species varied from 0.5% to 4.68%. The resulting MP tree and NJ tree showed relatively congruent phylogenetic relationships among these species, except Ae. caudata. Particularly, Ae. speltoides was basal within the two trees. The paraphyletic relationships between Ae. speltoides and two species of Sect. Sitopsis, and between Ae. uniaristata and two species of Sect. Comopyrum were supported strongly. The ITS data suggest that currently recognized sections within Aegilops should be reconsidered.

Base Composition↗

PubMed2000

Evolution of parental ITS regions of nuclear rDNA in allopolyploid Aegilops (Poaceae) species.

The genus Aegilops comprises approximately 25 diploid, tetraploid and hexaploid species, in which the genome types of all allopolyploids involve either U or D genome, or both of them. The internal transcribed spacer (ITS) region of 18S-26S nuclear ribosomal DNA (rDNA) from 11 allopolyploid species and 7 related diploid species in the genus were directly sequenced by pooled PCR products. Phylogenetic analyses for tracing evolutionary patterns of parental rDNA in allopolyploid species were performed using the neighbor-joining method. The D genome involved tree included three clades (CC-DDCC, DDMM-DDMMSS-DDMMUU, and MM-MhMh-DDNN), but did not include Ae. squarrosa (DD). It indicated that the rDNA of ancestral D genome had been somewhat differentiated in allopolyploids. The U genome involved tree showed that the allopolyploids and their common ancestor, Ae. umbellulata, formed a clade, suggesting that rDNA in UUMM and UUSS genomes has been homogenizing toward that of ancestral U genome. The phylogenetic pattern of U genome based on ITS sequences also supported the "pivotal-differential" hypothesis.

DNA, Plant↗

PubMed2000

2,4,6-trinitrotoluene reduction by carbon monoxide dehydrogenase from Clostridium thermoaceticum.

Purified CO dehydrogenase (CODH) from Clostridium thermoaceticum catalyzed the transformation of 2,4,6-trinitrotoluene (TNT). The intermediates and reduced products of TNT transformation were separated and appear to be identical to the compounds formed by C. acetobutylicum, namely, 2-hydroxylamino-4,6-dinitrotoluene (2HA46DNT), 4-hydroxylamino-2,6-dinitrotoluene (4HA26DNT), 2, 4-dihydroxylamino-6-nitrotoluene (24DHANT), and the Bamberger rearrangement product of 2,4-dihydroxylamino-6-nitrotoluene. In the presence of saturating CO, CODH catalyzed the conversion of TNT to two monohydroxylamino derivatives (2HA46DNT and 4HA26DNT), with 4HA26DNT as the dominant isomer. These derivatives were then converted to 24DHANT, which slowly converted to the Bamberger rearrangement product. Apparent K(m) and k(cat) values of TNT reduction were 165 +/- 43 microM for TNT and 400 +/- 94 s(-1), respectively. Cyanide, an inhibitor for the CO/CO(2) oxidation/reduction activity of CODH, inhibited the TNT degradation activity of CODH.

Aldehyde Oxidoreductases↗

PubMed2000

Downregulation of major histocompatibility complex class I molecules by Kaposi's sarcoma-associated herpesvirus K3 and K5 proteins.

The T-cell-mediated immune response plays a central role in the defense against intracellular pathogens. To avoid this immune response, viruses have evolved elaborate mechanisms that target and modulate many different aspects of the host's immune system. A target common to many of these viruses is the major histocompatibility complex (MHC) class I molecules. Kaposi's sarcoma-associated herpesvirus (KSHV) encodes K3 and K5 zinc finger membrane proteins which remove MHC class I molecules from the cell surface. K3 and K5 exhibit 40% amino acid identity to each other and localize primarily near the plasma membrane. While K3 and K5 dramatically downregulated class I molecules, they displayed different specificities in downregulation of HLA allotypes. K5 significantly downregulated HLA-A and -B and downregulated HLA-C only weakly, but not HLA-E, whereas K3 downregulated all four HLA allotypes. This selective downregulation of HLA allotypes by K5 was partly due to differences in amino acid sequences in their transmembrane regions. Biochemical analyses demonstrated that while K3 and K5 did not affect expression and intracellular transport of class I molecules, their expression induced rapid endocytosis of the molecules. These results demonstrate that KSHV has evolved a novel immune evasion mechanism by harboring similar but distinct genes, K3 and K5, which target MHC class I molecules in different ways.

Animals↗