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Biomedical subjects

J Yu

Publications and source records attributed to J Yu.

At least 325 records · Page 18Linked to original sources
PubMed1999

Synthesis and MAO-B substrate properties of 1-methyl-4-heteroaryl-1,2,3,6-tetrahydropyridines.

The parkinsonian inducing drug 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is bioactivated in a reaction catalyzed by the flavoenzyme monoamine oxidase B (MAO-B) to form the corresponding dihydropyridinium and subsequently pyridinium metabolites. As part of our ongoing studies to characterize the structural features responsible for this unexpected biotransformation, we have examined the MAO-B substrate properties of a variety of MPTP analogues bearing various heteroaryl groups at the 4-position of the tetrahydropyridinyl ring. The newly synthesized analogues are 4-(1-methylimidazol-2-yl)-, 4-(3-methylfuran-2-yl)-, 4-(3-methylthien-2-yl)-, 4-(3,4-dimethylpyrrol-1-yl)-, 4-(3-methylpyrrol-2-yl)-, and 4-(1,3-dimethylpyrrol-2-yl)-1-methyl-1,2,3,6-tetrahydropyridine. Except for the 4-(1-methylimidazol-2-yl) analogue, all compounds displayed good to excellent substrate properties. The 1-methyl-4-(3-methylfuran-2-yl) analogue is the most active member of this series with a kcat/Km value greater than 8,500 min(-1)mM(-1). The results of these studies are discussed in terms of catalytic pathways proposed for MAO-B.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1999

Synthesis and monoamine oxidase B substrate properties of 1-methyl-4-heteroaryl-1,2,3,6-tetrahydropyridines.

Six analogues of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine [MPTP, (1)] bearing various heteroaryl groups at C-4 were synthesized and examined for their monoamine oxidase B substrate properties. The C-4 substituents include the 1-ethylpyrrol-2-yl, 1-propylpyrrol-2-yl, 1-isopropylpyrrol-2-yl, 1-cyclopropylpyrrol-2-yl, 3-ethylfuran-2-yl and 3-ethylthien-2-yl groups. The results provide information concerning steric and polar interactions between the C-4 substituent and the active site of MAO-B that are transmitted to the position of oxidation at C-6 of the tetrahydropyridinyl moiety.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1999

DsbA: a protein-folding catalyst contributing to bacterial virulence.

DsbA, a periplasmic thiol:disulphide oxidoreductase, catalyses the folding of various factors, among which are virulence determinants or the components of type III secretory machinery. It is also necessary for intracellular survival and cell-to-cell spread of the intracellular pathogen Shigella flexneri.

Animals↗
PubMed1999

Repressor-AFLR interaction modulates aflatoxin biosynthesis in Aspergillus parasiticus.

Regulation of aflatoxin (AF) biosynthesis likely involves a complex interplay of positive- and negative-acting factors that are affected by physiological cues responsive to internal and external stimuli. These factors, presumably, modulate the expression of the AF pathway-specific regulatory gene, aflR, whose product, AFLR, a zinc cluster transcription factor, then turns on or off the transcription of other AF genes. To determine if the AFLR carboxyl region (AFLRC) interacts with positive- or negative-acting proteins, we fused the Aspergillus parasiticus aflR carboxyl coding region (aflRC) to the promoter of A. parasiticus nitrite reductase gene (niiA(p)::aflRC), and transformed it into A. parasiticus SRRC 2043. Transformants that contained two copies of niiA(p)::aflRC, one at the niaD locus and another at the aflR locus, overproduced AF precursors independent of the nitrogen source. The higher copy number of the integrated niiA(p)::aflRC correlated with increased production of AF precursors by the transformants as well as increased expression of both aflRC and native aflR in potato dextrose broth and A&M medium. Since aflRC does not encode a DNA-binding domain, the expressed AFLRC should not bind to the promoters of AF pathway genes and affect transcription directly. The results are consistent with AFLRC titrating out a putative repressor that interacts with AFLR under different growth conditions and modulates AF biosynthesis. This interaction also indirectly affects sclerotial development.

Aflatoxins↗
PubMed1999

The components of an inorganic physiological buffer for Penaeus chinensis.

Shrimp Penaeus chinensis, 5--8 cm in body-length were held in laboratory aquaria after collection from culture ponds at Laoshan in Qingdao in 1992. The osmotic pressure and the concentration of inorganic components of hemolymph in the shrimp were measured. The optimum pH and osmolarity of the buffer were determined from tests on primary cultures of hepatopancreatocytes in vitro. The results indicate that the hemolymph is homoiosmotic to 27 ppt seawater at 870 mOsm/L; osmotic regulation of shrimp is strong in low-salinity seawater, but weak in high-salinity seawater; the concentration of Mg(2+) ions in hemolymph is much less than that in seawater, while [K(+)] and [Ca(2+)] are much higher; hepatopancreatocytes in vitro have optimal survival at osmolarity range of 780-1, 100 mOsm/L and pH 6.5. Accordingly, an inorganic physiological buffer for P. chinensis (PPB, Penaeid Physiological Buffer) was prepared and contains NaCl, 23.0 g/L; K(2)SO(4), 1.1 g/L; CaCl(2), 1.6 g/L; MgSO(4).7H(2)O, 1.6 g/L; NaH(2)PO(4).2H(2)O, 0.35 g/L; NaHCO(3), 0. 05 g/L; pH 6.5; osmolarity, 867.9 mOsm/L. For storage of PPB, 4 stock buffers are made: 1) 10x PPB-A, which contains 230 g/L NaCl, 11 g/L K(2)SO(4), and 3.5 g/L NaH(2)PO(4).2H(2)O; 2) 40x PPB-B, which contains 64 g/L MgSO(4).7H(2)O; 3) 40x PPB-C, which contains 64 g/L CaCl(2); and 4) 100x PPB-D, which contains 50 g/L NaHCO(2). In order to increase the buffering capacity, PPB may be modified (to His-PPB), in which 5 g/L Histidine.HCl is used as the buffer reagent in place of NaH(2)PO(4) and NaHCO(3).

Animals↗
PubMed1999

c-MYC interacts with INI1/hSNF5 and requires the SWI/SNF complex for transactivation function.

Chromatin organization plays a key role in the regulation of gene expression. The evolutionarily conserved SWI/SNF complex is one of several multiprotein complexes that activate transcription by remodelling chromatin in an ATP-dependent manner. SWI2/SNF2 is an ATPase whose homologues, BRG1 and hBRM, mediate cell-cycle arrest; the SNF5 homologue, INI1/hSNF5, appears to be a tumour suppressor. A search for INI1-interacting proteins using the two-hybrid system led to the isolation of c-MYC, a transactivator. The c-MYC-INI1 interaction was observed both in vitro and in vivo. The c-MYC basic helix-loop-helix (bHLH) and leucine zipper (Zip) domains and the INI1 repeat 1 (Rpt1) region were required for this interaction. c-MYC-mediated transactivation was inhibited by a deletion fragment of INI1 and the ATPase mutant of BRG1/hSNF2 in a dominant-negative manner contingent upon the presence of the c-MYC bHLH-Zip domain. Our results suggest that the SWI/SNF complex is necessary for c-MYC-mediated transactivation and that the c-MYC-INI1 interaction helps recruit the complex.

Binding Sites↗
PubMed1999

Protection of human breast cancer cells from complement-mediated lysis by expression of heterologous CD59.

CD59, decay accelerating factor (DAF) and membrane cofactor protein (MCP) are widely expressed cell surface glycoproteins that protect host cells from the effects of homologous complement attack. Complement inhibitory activity of these proteins is species-selective. We show that the human breast cancer cell line MCF7 is relatively resistant to lysis by human complement, but is effectively lysed by rat or mouse complement. CD59, DAF and MCP were all shown to be expressed by MCF7. The species-selective nature of CD59 activity was used to demonstrate directly the effectiveness of CD59 at protecting cancer cells from complement-mediated lysis. cDNAs encoding rat and mouse CD59 were separately transfected into MCF7 cells, and cell populations expressing high levels of the rodent CD59 were isolated by cell sorting. Data show that rat and mouse CD59 were highly effective at protecting transfected MCF7 cells from lysis by rat and mouse complement, respectively. Data further reveal that rat CD59 is not effective against mouse complement, whereas mouse CD59 is effective against both mouse and rat complement. These studies establish a model system for relevant in vivo studies aimed at determining the effect of complement regulation on tumourigenesis, and show that for effective immunotherapy using complement-activating anti-tumour antibodies, the neutralization of CD59 and/or other complement inhibitory molecules will probably be required.

Animals↗
PubMed1999

Involvement of the exopolysaccharide alginate in the virulence and epiphytic fitness of Pseudomonas syringae pv. syringae.

Alginate, a co-polymer of O-acetylated beta-1,4-linked D-mannuronic acid and L-guluronic acid, has been reported to function in the virulence of Pseudomonas syringae, although genetic studies to test this hypothesis have not been undertaken previously. In the present study, we used a genetic approach to evaluate the role of alginate in the pathogenicity of P. syringae pv. syringae 3525, which causes bacterial brown spot on beans. Alginate biosynthesis in strain 3525 was disrupted by recombining Tn5 into algL, which encodes alginate lyase, resulting in 3525.L. Alginate production in 3525.L was restored by the introduction of pSK2 or pAD4033, which contain the alginate biosynthetic gene cluster from P. syringae pv. syringae FF5 or the algA gene from P. aeruginosa respectively. The role of alginate in the epiphytic fitness of strain 3525 was assessed by monitoring the populations of 3525 and 3525.L on tomato, which is not a host for this pathogen. The mutant 3525.L was significantly impaired in its ability to colonize tomato leaves compared with 3525, indicating that alginate functions in the survival of strain 3525 on leaf surfaces. The contribution of alginate to the virulence of strain 3525 was evaluated by comparing the population dynamics and symptom development of 3525 and 3525.L in bean leaves. Although 3525. L retained the ability to form lesions on bean leaves, symptoms were less severe, and the population was significantly reduced in comparison with 3525. These results indicate that alginate contributes to the virulence of P. syringae pv. syringae 3525, perhaps by facilitating colonization or dissemination of the bacterium in planta.

Adaptation, Biological↗
PubMed1999

The predictive value of white cell or CD34+ cell count in the peripheral blood for timing apheresis and maximizing yield.

BACKGROUND: The collection of peripheral blood stem and progenitor cells (PBPCs) for transplantation can be time-consuming and expensive. Thus, the utility of counting CD34+ cells and white cells (WBCs) in the peripheral blood was evaluated as a predictor of CD34+ cell yield in the apheresis component. STUDY DESIGN AND METHODS: The WBC and CD34+ cell counts in the peripheral blood and the apheresis components from 216 collections were assessed. Sixty-three patients underwent mobilization with chemotherapy plus filgrastim, and 17 patients and 14 allogeneic PBPC donors did so with filgrastim alone. The relationship between the number of WBC and CD34+ cells in the peripheral blood and in the apheresis component was analyzed by using rank correlation and linear regression analysis. RESULTS: The correlation coefficient for CD34+ cells per liter of peripheral blood with CD34+ cell yield (x 10(6)/kg) was 0.87 (n = 216 collections). This correlation existed for many patient and collection variables. However, patients with acute myeloid leukemia had fewer CD34+ cells in the apheresis component at any level of peripheral blood CD34+ cell count. Components collected from patients with CD34+ cell counts below 10 x 10(6) per L in the peripheral blood contained a median of 0.75 x 10(6) CD34+ cells per kg. When the WBC count in the blood was below 5.0 x 10(9) per L, the median number of CD34+ cells in the peripheral blood was 5.6 x 10(6) per L (range, 1.0-15.5 x 10(6)/L). A very poor correlation was found between the WBC count in the blood and the CD34+ cell yield (p = 0.12, n = 158 collections). CONCLUSION: The number of CD34+ cells, but not WBCs, in the peripheral blood can be used as a predictor for timing of apheresis and estimating PBPC yield. This is a robust relationship not affected by a variety of patient and collection factors except the diagnosis of acute myeloid leukemia. Patients who undergo mobilization with chemotherapy and filgrastim also should undergo monitoring of peripheral blood CD34+ cell counts, beginning when the WBC count in the blood exceeds 1.0 to 5.0 x 10(9) per L.

Adolescent↗
PubMed1999

Effects of genistein and daidzein on membrane characteristics of HCT cells.

Genistein and daidzein are two major isoflavonoids in soybeans. They have received increasing attention because of their possible role in cancer prevention. In the present investigation, the human colon tumor (HCT) cell line was used to investigate the effect of isoflavonoids on cell growth with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide colorimetric assay. We found that genistein and/or daidzein could inhibit growth in HCT cells. The 50% inhibitory concentrations of genistein and daidzein were 15 and 40 microM, respectively. Fluorescent polarization, quasielastic light scattering, and circular dichroism were used to study the influence of isoflavonoids on membrane characteristics of HCT cells, including membrane fluidity, density of cell surface charge, and membrane protein conformation. Membrane fluidity of HCT cells was obviously reduced by genistein, but not by daidzein. The effect of genistein was time and dose dependent. In addition, genistein and daidzein could reduce the density of cell surface charge and increase the order of membrane protein conformation. All these changes may represent one of the mechanisms of the effect of isoflavonoids on growth inhibition, differentiation promotion, and transfer interference in a tumor cell line.

Anticarcinogenic Agents↗
PubMed1999

Poloxamine 1107 sealing of radiopermeabilized erythrocyte membranes.

PURPOSE: Lipid peroxidation-mediated permeabilization of cell membranes following intense ionizing irradiation is well documented. This form of membrane radiopermeabilization leads to rapid exhaustion of cellular high-energy compounds, resulting in the acute onset of cellular necrosis. Strategies to reverse the process of necrosis and preserve cell viability require membrane sealing. This report documents the relative efficacy of Poloxamine 1107, a non-ionic surfactant, compared with other polymers, in sealing radiopermeabilized cell membranes. MATERIALS AND METHODS: Isolated erythrocytes were exposed to 600 Gy 60Co irradiation at a dose rate of 1.3 Gy/s. Different polymer compounds were added 10 min later to the irradiated cell suspensions. At 2 h later the haemoglobin content in the supernatants was determined spectrophotometrically. RESULTS: Compared with the non-treated irradiated control, Poloxamine 1107 significantly reduced the leakage of haemoglobin from irradiated erythrocytes. Poloxamer 188 and dextran at equal concentrations had no significant reverse effect on the irradiation-mediated increased membrane permeability. The amount of haemoglobin released from irradiated erythrocytes was inversely related to the Poloxamine 1107 concentration. CONCLUSIONS: This study demonstrates the capability of Poloxamine 1107 to seal radiopermeabilized cell membranes. Thus, surfactants such as Poloxamine 1107 might be useful as a therapeutic agent in the treatment of high-dose radiation injuries since cellular necrosis due to metabolic exhaustion following radiopermeabilization of their membranes might be prevented.

Cell Membrane Permeability↗
PubMed1999

Effects of chlorophyll availability on phycobilisomes in Synechocystis sp. PCC 6803.

Inactivation of the chlL gene in Synechocystis sp. PCC 6803 resulted in negligible chlorophyll content when the mutant was grown in darkness. Upon phycocyanin excitation at 580 nm, the 77K fluorescence spectrum of dark-grown cells showed three peaks at 648 nm, 665 nm, and 685 nm, this last being the largest. This reflects the functional presence of major components of phycobilisomes, including phycocyanin, allophycocyanin, and the terminal emitter, and efficient energy transfer between these components. As expected, no fluorescence emission peaks corresponding to chlorophyll in the photosystems were observed. Intact phycobilisomes could be isolated from the dark-grown chlL-deletion mutant. However, the phycobilisomes had a lower efficiency of energy transfer than did those isolated from the light-grown mutant, probably because of a decreased phycobilisome stability in the absence of chlorophyll. Exposing the dark-grown chlL-deletion mutant to light triggered the biosynthesis of chlorophyll. For the first 6 h in the light, upon phycocyanin excitation at 580 nm, the 77K fluorescence emission spectrum of greening cells was identical to that of dark-grown cells that lacked significant amounts of chlorophyll. With increased chlorophyll synthesis, gradual energy transfer from phycobilisomes to the two photosystems can be demonstrated.

Bacterial Proteins↗
PubMed1999

Enhancement of spectral analysis of myoelectric signals during static contractions using wavelet methods.

In this paper, we introduce wavelet packets as an alternative method for spectral analysis of surface myoelectric (ME) signals. Both computer synthesized and real ME signals are used to investigate the performance. Our simulation results show that wavelet packet estimate has slightly less mean square error (MSE) than Fourier method, and both methods perform similarly on the real data. Moreover, wavelet packets give us some advantages over the traditional methods such as multiresolution of frequency, as well as its potential use for effecting time-frequency decomposition of the nonstationary signals such as the ME signals during dynamic contractions. We also introduce wavelet shrinkage method for improving spectral estimates by significantly reducing the MSE's for both Fourier and wavelet packet methods.

Action Potentials↗
PubMed1999

The carboxy-terminal portion of the aflatoxin pathway regulatory protein AFLR of Aspergillus parasiticus activates GAL1::lacZ gene expression in Saccharomyces cerevisiae.

AFLR, a DNA-binding protein of 444 amino acids, transactivates the expression of aflatoxin biosynthesis genes in Aspergillus parasiticus and Aspergillus flavus, as well as the sterigmatocystin synthesis genes in Aspergillus nidulans. We show here by fusion of various aflR coding regions to the GAL4 DNA-binding coding region that the AFLR carboxyl terminus contained a region that activated GAL1::lacZ gene expression in Saccharomyces cerevisiae and that the AFLR internal region was required for the activation activity. Compared to the AFLR carboxy-terminal fusion protein (AFLRC), a mutant AFLRC retained approximately 75% of the activation activity after deletion of three acidic amino acids, Asp365, Glu366, and Glu367, in a previously identified acidic stretch. Removal of the carboxy-terminal amino acid, Glu444, did not affect the activation activity. Substitutions of acidic Glu423, Asp439, or Asp436/Asp439 with basic amino acids, Lys and His, resulted in 10- to 15-fold-lower activation activities. Strikingly, the Asp436His mutation abolished the activation activity. Substitutions of basic His428 and His442 with acidic Asp resulted in 20 and 40% decreases in the activation activities, respectively. Simultaneous substitutions of Arg427, Arg429, and Arg431 with Leu also significantly decreased the activation activity; the decrease was approximately 50-fold. Results suggest that the AFLR carboxy-terminal region is involved in transcription activation and that total acidity in this region is not a major determinant of AFLR's activation ability in S. cerevisiae.

Amino Acid Sequence↗
PubMed1999

The cloned locus of enterocyte effacement from enterohemorrhagic Escherichia coli O157:H7 is unable to confer the attaching and effacing phenotype upon E. coli K-12.

The locus of enterocyte effacement (LEE) pathogenicity island of enterohemorrhagic Escherichia coli (EHEC) O157:H7 possesses the same genes in identical order and orientation as the LEE of enteropathogenic E. coli (EPEC) O127:H6 but is unable to form attaching and effacing (A/E) lesions or to secrete Esp proteins when it is cloned in an E. coli K-12 background. The A/E phenotype could not be restored by trans complementation with a variety of cloned EPEC LEE fragments, suggesting functional and/or regulatory differences between the LEE pathogenicity islands of EPEC O127:H6 and EHEC O157:H7.

Chromosome Mapping↗
PubMed1999

Signals from the Ras, Rac, and Rho GTPases converge on the Pak protein kinase in Rat-1 fibroblasts.

Ras plays a key role in regulating cellular proliferation, differentiation, and transformation. Raf is the major effector of Ras in the Ras > Raf > Mek > extracellular signal-activated kinase (ERK) cascade. A second effector is phosphoinositide 3-OH kinase (PI 3-kinase), which, in turn, activates the small G protein Rac. Rac also has multiple effectors, one of which is the serine threonine kinase Pak (p65(Pak)). Here we show that Ras, but not Raf, activates Pak1 in cotransfection assays of Rat-1 cells but not NIH 3T3 cells. We tested agents that activate or block specific components downstream of Ras and demonstrate a Ras > PI 3-kinase > Rac/Cdc42 > Pak signal. Although these studies suggest that the signal from Ras through PI 3-kinase is sufficient to activate Pak, additional studies suggested that other effectors contribute to Pak activation. RasV12S35 and RasV12G37, two effector mutant proteins which fail to activate PI 3-kinase, did not activate Pak when tested alone but activated Pak when they were cotransfected. Similarly, RacV12H40, an effector mutant that does not bind Pak, and Rho both cooperated with Raf to activate Pak. A dominant negative Rho mutant also inhibited Ras activation of Pak. All combinations of Rac/Raf and Ras/Raf and Rho/Raf effector mutants that transform cells cooperatively stimulated ERK. Cooperation was Pak dependent, since all combinations were inhibited by kinase-deficient Pak mutants in both transformation assays and ERK activation assays. These data suggest that other Ras effectors can collaborate with PI 3-kinase and with each other to activate Pak. Furthermore, the strong correlation between Pak activation and cooperative transformation suggests that Pak activation is necessary, although not sufficient, for cooperative transformation of Rat-1 fibroblasts by Ras, Rac, and Rho.

3T3 Cells↗