Biomedical subjects

Z Qu

Publications and source records attributed to Z Qu.

At least 19 recordsLinked to original sources

PubMed2002

Muscle-derived cell-mediated ex vivo gene therapy for urological dysfunction.

We have tested the feasibility of muscle-based gene therapy and tissue engineering for urological dysfunction using highly purified muscle-derived cells (MDC) that display stem cell characteristics. We then explored the potential use of these MDC as an alternative therapy for the treatment of impaired detrusor contractility. The MDC were genetically engineered to express the gene encoding beta-galactosidase and injected into the bladder walls of SCID mice. The injected bladders were harvested at various time-points after injection and assayed for beta-galactosidase activity; the presence of myofibers within the injected tissue was determined by detection of fast myosin heavy chain isoform (MyHCs). We have demonstrated that the injected MDC are capable of not only surviving in the lower urinary tract, but also improving the contractility of the bladder following an induced injury. Two potential mechanisms can be used to explain this finding. First, we have observed that some of the beta-galactosidase-expressing cells expressed alpha-smooth muscle actin, suggesting a differentiation into smooth muscle. Second, a stain for acetylcholine receptors (AChRs), which identifies the location of neuromuscular junctions, revealed that the myofibers derived from the doner cells became innervated into the bladder as early as 2 weeks after injection. These results suggest that gene therapy and tissue engineering based on MDC potentially can be used for urological dysfunction.

Actins↗

PubMed2001

A kinetic study on the pairwise competition reaction of alpha-diazo esters with rhodium(II) catalysts: implication for the mechanism of Rh(II)-carbene transfer.

The relative rate constants for the Rh(II)-mediated diazo decomposition of a series of para- or meta-substituted diazophenylacetates were measured through intermolecular competition. The kinetic data were further subjected to Hammett correlation analysis and were found to have better linear correlation with sigma(+). Reaction constants for four Rh(II) catalysts have been obtained, Rh(2)(OAc)(4) (rho = -1.29 with sigma(+), r = -0.99), Rh(2)(Ooct)(4) (rho = -1.31 with sigma(+), r = -0.99), Rh(2)(acam)(4) (rho = -1.18 with sigma(+), r = -0.99), Rh(2)(O(2)CCF(3))(4) (rho = -1.46 with sigma(+), r = -0.99). The mechanistic implications of these data are discussed.

Journal Article↗

PubMed2001

Mechanisms of ascorbic acid recycling in human erythrocytes.

Vitamin C, or ascorbic acid, is efficiently recycled from its oxidized forms by human erythrocytes. In this work the dependence of this recycling on reduced glutathione (GSH) was evaluated with regard to activation of the pentose cycle and to changes in pyridine nucleotide concentrations. The two-electron-oxidized form of ascorbic acid, dehydroascorbic acid (DHA) was rapidly taken up by erythrocytes and reduced to ascorbate, which reached intracellular concentrations as high as 2 mM. In the absence of D-glucose, DHA caused dose-dependent decreases in erythrocyte GSH, NADPH, and NADH concentrations. In the presence of 5 mM D-glucose, GSH and NADH concentrations were maintained, but those of NADPH decreased. Reduction of extracellular ferricyanide by erythrocytes, which reflects intracellular ascorbate recycling, was also enhanced by D-glucose, and ferricyanide activated the pentose cycle. Diethylmaleate at concentrations up to 1 mM was found to specifically deplete erythrocyte GSH by 75-90% without causing oxidant stress in the cells. Such GSH-depleted erythrocytes showed parallel decreases in their ability to take up and reduce DHA to ascorbate, and to reduce extracellular ferricyanide. These results show that DHA reduction involves GSH-dependent activation of D-glucose metabolism in the pentose cycle, but that in the absence of D-glucose DHA reduction can also utilize NADH.

Ascorbic Acid↗

PubMed2001

Requirement for GSH in recycling of ascorbic acid in endothelial cells.

Ascorbic acid may be involved in the defense against oxidant stress in endothelial cells. Such a role requires that the cells effectively recycle the vitamin from its oxidized forms. In this work, we studied the ability of cultured bovine aortic endothelial cells (BAECs) to take up and reduce dehydroascorbic acid (DHA) to ascorbate, as well as the dependence of ascorbate recycling on intracellular GSH. BAECs took up and reduced DHA to ascorbate much more readily than they took up ascorbate. Although BAECs in culture did not contain ascorbate, ascorbate accumulated to concentrations of 2-3 mM in BAECs following incubation with 400 microM DHA. Extracellular ferricyanide oxidized intracellular ascorbate, which was recycled by the cells. Reduction of DHA, either when added to the cells or when generated in response to ferricyanide, caused significant decreases in intracellular GSH concentrations. Depletion of intracellular GSH with 1-chloro-2,4-dinitrobenzene, diethylmaleate, and diamide almost abolished the ability of the cells to reduce DHA to ascorbate. DHA reduction by thioredoxin reductase was evident in dialyzed cell extracts, but occurred at rates far lower than direct GSH reduction of DHA. These results suggest that maximal rates of DHA reduction, and thus recycling of ascorbate from DHA, are dependent upon GSH in these cells.

Animals↗

PubMed2001

1,2-aryl and 1,2-hydride migration in transition metal complex catalyzed diazo decomposition: a novel approach to alpha-aryl-beta-enamino esters.

N-Tosyl diazoketamines were prepared by addition of the ethyl alpha-diazoacetate anion to N-sulfonylimines. The diazo decomposition of the diazoketamines with Rh(2)(OAc)(4) complex resulted in aryl migration to give alpha-aryl-beta-enamino esters in good yields and high stereoselectivity. The effect of the catalysts on the migratory aptitude of 1,2-aryl over 1,2-hydride migration was studied. A reaction mechanism involving a "bridged" phenonium ion is proposed. Reaction: see text.

Journal Article↗

PubMed2001

Recycling of the ascorbate free radical by human erythrocyte membranes.

Reduction of the ascorbate free radical (AFR) at the plasma membrane provides an efficient mechanism to preserve the vitamin in a location where it can recycle alpha-tocopherol and thus prevent lipid peroxidation. Erythrocyte ghost membranes have been shown to oxidize NADH in the presence of the AFR. We report that this activity derives from an AFR reductase because it spares ascorbate from oxidation by ascorbate oxidase, and because ghost membranes decrease steady-state concentrations of the AFR in a protein- and NADH-dependent manner. The AFR reductase has a high apparent affinity for both NADH and the AFR (< 2 microM). When measured in open ghosts, the reductase is comprised of an inner membrane activity (both substrate sites on the cytosolic membrane face) and a trans-membrane activity that mediates extracellular AFR reduction using intracellular NADH. However, the trans-membrane activity constitutes only about 12% of the total measured in ghosts. Ghost AFR reductase activity can also be differentiated from NADH-dependent ferricyanide reductase(s) by its sensitivity to the detergent Triton X-100 and insensitivity to enzymatic digestion with cathepsin D. This NADH-dependent AFR reductase could serve to recycle ascorbic acid at a crucial site on the inner face of the plasma membrane.

Ascorbic Acid↗

PubMed2001

Functional geometry of the permeation pathway of Ca2+-activated Cl-channels inferred from analysis of voltage-dependent block.

We examined the voltage-dependent block of Ca(2+)-activated Cl(-) channels by anthacene-9-carboxylic acid (A9C), diphenylamine-2-carboxylic acid (DPC), 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), and niflumic acid (NFA) in excised inside-out and outside-out patches from Xenopus oocytes. The fraction of the voltage field (delta) experienced by the blocking drug was determined from the voltage dependence of block. All the drugs blocked by entering the channel from the outside. delta was 0.6 for A9C, 0.3 for DPC and DIDS, and <0.1 for NFA. Because the voltage dependence of the drugs differed, the order of potency was also voltage-dependent. At +100 mV the order of potency was NFA > A9C > DIDS > DPC (K(i) (microm) = 10.1, 18.3, 48, and 111, respectively). Because the drugs are hydrophobic, they can cross the bilayer when applied from the inside and block the channel from the outside. The equilibrium geometries of the blockers were determined by molecular modeling and compared with their blocking positions (delta). This analysis suggests that the channel is an elliptical cone with the largest opening facing the extracellular space. The selectivity filter has an apparent size of 0.33 x 0.75 nm, because C(CN)(3)-, which has these dimensions, permeates. The external opening is at least 0.60 x 0.94 nm, because DPC has these dimensions and penetrates the channel approximately 30%.

Animals↗

PubMed2001

Coexistence of multiple spiral waves with independent frequencies in a heterogeneous excitable medium.

We studied the interactions and coexistence of stable spiral waves with independent frequencies in a heterogeneous excitable medium, using numerical simulations of a spatial system based on the FitzHugh-Nagumo cell model. When the heterogeneity of the medium exceeded a critical value, a transition took place from a single dominant spiral wave to a coexistence of multiple spiral waves with independent frequencies and n:n-1 wave conduction blocks. In this case, multiple spiral waves could coexist because they are "insulated" from each other by chaotic regions.

Biological Clocks↗

PubMed2001

Cloning, molecular characterization, and application of rice epiphytic Bacillus pumilus promoter fragments.

To establish a constitutive, high-efficiency expression system for Bacillus pumilus (B.P), we cloned random chromosomal DNA into promoter probe shuttle vector ECE7 and selected for strong promoter activity by chloramphenicol resistance of transformed B. pumilus cells. The nucleotide sequences of nine chromosomal fragments were determined. These DNA fragments range from 300 to 2200 bp in size. The transcription strength of these promoters was estimated by determination of CAT enzyme production in both E. coli and B. pumilus. Transcription start (TS) sites of the cat mRNA were located by primer extension by using total RNA. Preliminary analysis showed that three of the promoter sequences contain -35 and -10 regions like E. coli RNA polymerase sigma70 and B. subtilis sigma43 consensus sequences. One is similar to B. subtilis sigma29, the other two have no conserved sequences like any of the typical consensus sequences of the known sigma factors so far. To estimate the feasibility of the utilization of these promoters, one promoter fragment was subcloned and used to drive the expression of green fluorescent protein (GFP) in B. pumilus cells. This is the first report of B. pumilus promoters randomly cloning from total DNA and molecular analysis of their consensus sequences.

Bacillus↗

PubMed2001

Muscle-derived cell transplantation and differentiation into lower urinary tract smooth muscle.

OBJECTIVES: To explore the feasibility of primary skeletal muscle-derived cell (MDC)-based tissue engineering and gene transfer into the lower urinary tract and to explore whether the injected primary skeletal MDCs can persist and differentiate into myotubes and myofibers in the bladder wall. METHODS: Primary MDCs isolated from normal mice were first transduced with adenovirus encoding the expression of the beta-galactosidase reporter gene. Adult severe combined immunodeficiency mice (n = 12) were used in this study. The MDCs were injected into the right and left lateral bladder walls with a 10-microL Hamilton microsyringe. The amount of injected MDCs ranged from 1 to 1.5 x 10(6) cells. The tissue was harvested after 5, 35, and 70 days, sectioned, stained for fast myosin heavy chain, and assayed for beta-galactosidase expression. RESULTS: We observed a large number of cells expressing beta-galactosidase in the bladder wall at each time point. Many myotubes and myofibers expressing beta-galactosidase and positively stained for fast myosin heavy chain were also seen in the bladder wall at 35 and 70 days after injection. Additionally, the size of the injected MDCs significantly increased during the course of the study (P <0.05). CONCLUSIONS: We have demonstrated the long-term survival and beta-galactosidase expression of MDCs injected into the bladder wall. Moreover, our results suggest that some injected MDCs can differentiate into myofibers. These results suggest that MDCs can be a desirable substance for tissue engineering and an ex vivo method for gene transfer into the lower urinary tract.

Animals↗

PubMed2001

Impaired prostate tumorigenesis in Egr1-deficient mice.

The transcription factor early growth response protein 1 (EGR1) is overexpressed in a majority of human prostate cancers and is implicated in the regulation of several genes important for prostate tumor progression. Here we have assessed the effect of Egr1 deficiency on tumor development in two transgenic mouse models of prostate cancer (CR2-T-Ag and TRAMP). Using a combination of high-resolution magnetic resonance imaging and histopathological and survival analyses, we show that tumor progression was significantly impaired in Egr1-/- mice. Tumor initiation and tumor growth rate were not affected by the lack of Egr1; however, Egr1 deficiency significantly delayed the progression from prostatic intra-epithelial neoplasia to invasive carcinoma. These results indicate a unique role for Egr1 in regulating the transition from localized, carcinoma in situ to invasive carcinoma.

Animals↗

PubMed2001

Effect of anti-IL-6 and anti-10 monoclonal antibodies on the suppression of the normal T lymphocyte mitogenic response by steady state sickle cell disease sera.

Previously published work has shown that sera from healthy sickle cell disease (SCD) patients inhibits normal lymphocyte mitogenic response to phytohemagglutinin (PHA) in vitro. The current study is to attempt to ascertain what effect antibody to type 2 cytokines, interleukin (IL)-6 and 10, have on the suppression of lymphocyte PHA response by SCD sera. Peripheral blood mononuclear cells (PBMC), separated by density gradient were obtained from 2 healthy normal donors. Sera from 50 healthy SCD patients, 50 normal healthy controls and pooled normal O, Rh+ (O+) sera were utilized in standard in vitro PHA stimulation of PBMC cultures. Mitogenic responses were expressed as mean counts per minute (cpm) of triplicate cultures. Fifty triplicate cultures of PHA stimulated normal PBMC were done with 10% normal pooled O+, normal control and SCD steady state sera only. In addition 50 cultures were done with SCD sera containing 1 microg/ml of anti-IL-6 monoclonal antibody, as well as 28 SCD serum cultures containing 1 microg/ml of anti-IL-10 monoclonal antibody. The final 11 SCD serum culture experiments contained a combination of both anti-IL-6 and anti-IL-10 antibody, each at the concentration of 1 microg/ml. Results revealed > 15% suppression of mitogenic response in the SCD sera supplemented cultures as compared to control sera in 47/50 (94%) and in 40/50 (80%) of normal pooled O+, as calculated by mean cpm. The degree of suppression ranged from 17% to 98% in individual experiments. The addition of anti IL-6 antibody alone significantly improved mean cpm (> 20%) in 19/50 (38%) of SCD serum responses compared to O+ sera and 23/50 (46%) of control sera. Complete correction occurred in 9/50 (18%) of all SCD serum suppressions as compared to O+ sera and 6/50 (12%) when compared to control sera. Similarly, anti-IL-10 antibody decreased suppression of the mean cpm of SCD serum cultures in 18/28 (64%) and completely corrected 3/18 (11%). The combined antibody data revealed >20% increase in mean cpm in 10/11(91%) experiments. Inhibitors of mitogenic response were present in a significant percentage of the SCD sera utilized in the present study. The significant corrective effects of both monoclonal antibodies would seem to support the original hypothesis that high circulating levels of type 2 cytokines may represent the cell-mediated dependent inhibitory factors expressed in the sera of many healthy SCD patients.

Adolescent↗

PubMed2001

Persistence and survival of autologous muscle derived cells versus bovine collagen as potential treatment of stress urinary incontinence.

PURPOSE: We explored the use of autologous muscle derived cells as a method of treating stress urinary incontinence. We determined whether urethral muscle derived cell injection is feasible and compared it with bovine collagen injection. MATERIALS AND METHODS: Muscle derived cells isolated from female Sprague-Dawley rats were first transduced with retrovirus carrying the transgene for beta-galactosidase. We injected approximately 1 to 1.5 x 106 cells into the bladder wall and proximal urethra of 6 autologous animals. Tissue was harvested after 3 and 30 days, sectioned, stained for fast myosin heavy chain and assayed for beta-galactosidase. To compare muscle derived cell and bovine collagen injections 100 microl. of commercially available bovine collagen were also injected in Sprague-Dawley female rats. Tissue was harvested in 3 animals each after 3 and 30 days, sectioned and stained for trichrome. Subsequently, 3 adult SCID mice were used to compare the level of transgene expression at each time point after injecting 1.5 x 106 cells per injection, which were transduced with adenovirus carrying the transgene for beta-galactosidase. RESULTS: A large number of cells expressing beta-galactosidase were observed in the bladder and urethral wall 3 and 30 days after autologous cell injection in Sprague-Dawley rats. The persistence of primary muscle derived cells at 3 days was similar to that of collagen. However, at 30 days there was significant cell persistence while only a minimal amount of injected bovine collagen was detectable. Approximately 88% of the beta-galactosidase expression at day 3 remained at day 30 in SCID mice. CONCLUSIONS: We present 2 new findings important for the emerging field of urological tissue engineering, including the feasibility of injecting autologous skeletal muscle derived cells into the lower urinary tract and the greater persistence of such injected cells versus injected bovine collagen. Therefore, autologous muscle derived cell injection may be an attractive alternative treatment option for stress urinary incontinence.

Animals↗

PubMed2001

Electrophysiological heterogeneity and stability of reentry in simulated cardiac tissue.

Generation of wave break is a characteristic feature of cardiac fibrillation. In this study, we investigated how dynamic factors and fixed electrophysiological heterogeneity interact to promote wave break in simulated two-dimensional cardiac tissue, by using the Luo-Rudy (LR1) ventricular action potential model. The degree of dynamic instability of the action potential model was controlled by varying the maximal amplitude of the slow inward Ca(2+) current to produce spiral waves in homogeneous tissue that were either nearly stable, meandering, hypermeandering, or in breakup regimes. Fixed electrophysiological heterogeneity was modeled by randomly varying action potential duration over different spatial scales to create dispersion of refractoriness. We found that the degree of dispersion of refractoriness required to induce wave break decreased markedly as dynamic instability of the cardiac model increased. These findings suggest that reducing the dynamic instability of cardiac cells by interventions, such as decreasing the steepness of action potential duration restitution, may still have merit as an antifibrillatory strategy.

Action Potentials↗

PubMed2001

Effects of simulated ischemia on spiral wave stability.

Regional hyperkalemia during acute myocardial ischemia is a major factor promoting electrophysiological abnormalities leading to ventricular fibrillation (VF). However, steep action potential duration restitution, recently proposed to be a major determinant of VF, is typically decreased rather than increased by hyperkalemia and acute ischemia. To investigate this apparent contradiction, we simulated the effects of regional hyperkalemia and other ischemic components (anoxia and acidosis) on the stability of spiral wave reentry in simulated two-dimensional cardiac tissue by use of the Luo-Rudy ventricular action potential model. We found that the hyperkalemic "ischemic" area promotes wavebreak in the surrounding normal tissue by accelerating the rate of spiral wave reentry, even after the depolarized ischemic area itself has become unexcitable. Furthermore, wavebreak and fibrillation can be prevented if the dynamical instability of the normal tissue is reduced significantly by targeting electrical restitution properties, suggesting a novel therapeutic approach.

Action Potentials↗

PubMed2001

Patterns of wave break during ventricular fibrillation in isolated swine right ventricle.

Several different patterns of wave break have been described by mapping of the tissue surface during fibrillation. However, it is not clear whether these surface patterns are caused by multiple distinct mechanisms or by a single mechanism. To determine the mechanism by which wave breaks are generated during ventricular fibrillation, we conducted optical mapping studies and single cell transmembrane potential recording in six isolated swine right ventricles (RV). Among 763 episodes of wave break (0.75 times x s(-1) x cm(-2)), optical maps showed three patterns: 80% due to a wave front encountering the refractory wave back of another wave, 11.5% due to wave fronts passing perpendicular to each other, and 8.5% due to a new (target) wave arising just beyond the refractory tail of a previous wave. Computer simulations of scroll waves in three-dimensional tissue showed that these surface patterns could be attributed to two fundamental mechanisms: head-tail interactions and filament break. We conclude that during sustained ventricular fibrillation in swine RV, surface patterns of wave break are produced by two fundamental mechanisms: head-tail interaction between waves and filament break.

Action Potentials↗

PubMed2001

[Mapping the interaction site of Rpb2 and Rpb3 subunit of fission yeast RNA polymerase II].

To map the interacting site of subunit Rpb2 to subunit Rpb3 of RNA polymerase II in fission yeast Schizosaccharomyces pombe, the yeast two-hybrid system was employed in this paper to screen the interacting clones between Rpb2 and Rpb3.4 fragments of Rpb2 cDNA were cloned into the Ga14 BD vector pAS2. The 4 clones were named as pAS2 Rpb2-1, 2-2, 2-3 and 2-4, respectively. The complete cDNA of Rpb3 was cloned into the Gal 4 AD vector pGADGH. The clone was named as pGADGH Rpb3. The two-hybrid plasmids pGADGH Rpb3 and pAS2Rpb2-1, 2-2, 2-3 or 2-4 respectively were cotransformed into host cell yeast Y190. The interaction positive cotransformants were identified by beta-gal activity assay. The beta-gal positive cotransformants were selected from pGADGH Rpb3 and pAS2Rpb2-4 two-hybrid system. DNA sequencing and alignment results showed that the interacting site of Rpb2 to Rpb3 located within the fragment from base 2701 to 2966 of Rpb2 cDNA, or within the C-termini polypeptide from amino acid 902 to 989 of Rpb2 protein.

Amino Acid Sequence↗

PubMed2000

Ventricular fibrillation: how do we stop the waves from breaking?

Combined experimental and theoretical developments have demonstrated that in addition to preexisting electrophysiological heterogeneities, cardiac electrical restitution properties contribute to breakup of reentrant wavefronts during cardiac fibrillation. Developing therapies that favorably alter electrical restitution properties have promise as a new paradigm for preventing fibrillation.

Action Potentials↗