Biomedical subjects

J Yu

Publications and source records attributed to J Yu.

At least 163 records · Page 9Linked to original sources

PubMed2000

Do subplate neurons comprise a transient population of cells in developing neocortex of rats?

Studies were undertaken to determine whether neurons of the subplate layer represent a transient or stable population of cells in developing neocortex of rat. The first set of studies sought to determine the fraction of subplate neurons that is lost during early postnatal development. The optical dissector method was used to analyze fluorescently stained material in animals the age of postnatal day 0 (P0) to P40. These results demonstrate a reduction of slightly less than half of the total number of subplate neurons from P0 to P40. Counts of labeled cells in littermates at varied ages after [(3)H]thymidine or BRDU treatment on gestational day 14 (G14 - birthdate of occipital subplate neurons) or G18 (birthdate of layers III-IV neurons) demonstrate loss of approximately 50% of neurons in the subplate layer between P0 and P40, somewhat greater than the loss of neurons from cortical layers III-IV. The second set of studies investigated whether subplate neurons display cellular atrophy during postnatal development. Analysis of subplate neurons injected intracellularly with Lucifer yellow in fixed slice preparations indicates no reduction in soma size, number of dendrites, or extent of dendritic fields of subplate neurons taken from animals age P0 to P60. The third set of studies investigated whether functional markers of subplate neurons are reduced during postnatal development. Analysis of tissue stained histochemically for cytochrome oxidase or acetylcholinesterase, or stained immunocytochemically for GABA, somatostatin, or neuropeptide Y, demonstrate a remarkable loss of expression of staining patterns from late gestational ages to P20. These data demonstrate that, although subplate neurons seem not to be a transient population of cells in the usual sense of being eliminated by cell death or structural atrophy, the loss of histochemical and immunocytochemical markers indicates that they may be a functionally transient population of cells.

Acetylcholinesterase↗

PubMed2000

Design, synthesis, and X-ray crystal structure of a potent dual inhibitor of thymidylate synthase and dihydrofolate reductase as an antitumor agent.

A novel N-¿2-amino-4-methyl[(pyrrolo[2, 3-d]pyrimidin-5-yl)ethyl]benzoyl¿-L-glutamic acid (3a) was designed and synthesized as a potent dual inhibitor of thymidylate synthase (TS) and dihydrofolate reductase (DHFR) and as an antitumor agent. Compound 3b, the N7-benzylated analogue of 3a, was also synthesized as an antitumor agent. The synthesis of 3a was accomplished via a 12-step sequence which involved the synthesis of 2-amino-4-methylpyrrolo[2,3-d]pyrimidine (10) in 5 steps from 2-acetylbutyrolactone. Protection of the 2-amino group of 10 and regioselective iodination at the 5-position followed by palladium-catalyzed coupling afforded intermediate 14 which was converted to 3a by reduction and saponification. Similar synthetic methodology was used for 3b. X-ray crystal structure of the ternary complex of 3a, DHFR, and NADPH showed that the pyrrolo[2, 3-d]pyrimidine ring binds in a "2,4-diamino mode" in which the pyrrole nitrogen mimics the 4-amino moiety of 2,4-diaminopyrimidines. This is the first example of a classical pyrrolo[2,3-d]pyrimidine antifolate shown to have this alternate mode of binding to DHFR. Compounds 3a and 3b were more inhibitory than LY231514 against TS from Lactobacillus casei and Escherichia coli. Analogue 3a was also more inhibitory against DHFR from human, Toxoplasma gondii, and Pneumocystis carinii. Evaluation of 3a against methotrexate (MTX)-resistant cell lines with defined mechanisms indicated that cross-resistance of 3a was much lower than that of MTX. Metabolite protection studies and folylpoly-gamma-glutamate synthetase studies suggest that the antitumor activity of 3a against the growth of tumor cells in culture is a result of dual inhibition of TS and DHFR. Compound 3a inhibited the growth of CCRF-CEM and FaDu cells in culture at ED(50) values of 12.5 and 7.0 nM, respectively, and was more active against FaDu cells than MTX. In contrast, compound 3b was inactive against both cell lines. Compound 3a was evaluated in the National Cancer Institute in vitro preclinical antitumor screening program and afforded IG(50) values in the nanomolar range against a number of tumor cell lines.

Animals↗

PubMed2000

A method for determination of pyridine nucleotides using a single extract.

Intracellular redox levels play an important role in physiology and pathophysiology. The principal intracellular reductant is NADPH, which is required for both the proper activity of the entire antioxidant system and important prooxidant enzymes such as nitric oxide synthase and NADPH oxidase. Thus an easy and accurate measurement of NADPH is very desirable. The method described in this paper is based on the fact that NADH and NADPH (not NAD(+) and NADP(+)) affect absorbance at 340 nm. A single cell extract is separated into three aliquots (A(1), A(2), and A(3)). A(1) is untreated and the absorbance at 340 nm is measured. A(2) is treated with an enzyme that converts all of the NADP(+) to NADPH and then the absorbance at 340 nm is measured. A(3) is treated with an enzyme that converts all of the NADPH to NADP(+) and then the absorbance at 340 nm is measured. A(1) - A(3) is the NADPH content and A(2) - A(1) is the NADP(+) content of the extract. Using this method, we have obtained full recovery of all added nucleotides from cell extracts, thus making the method suitable for the quick determination of NADP(+) and NADPH in living cells. We conclude that this method for the measurement of NADP(+) and NADPH is rapid, simple, accurate, and reliable.

Animals↗

PubMed2000

Molecular lens of the nonresonant dipole force

A cylindrical molecular lens is formed by focusing a nanosecond IR laser pulse. Trajectories of a CS2 molecular beam deflected by the lens are traced using the velocity map imaging technique. The characteristic lens parameters including the focal length, minimum beam width, and distance to the minimum-width position are determined. The laser intensity dependence of the parameters is in good agreement with theoretical predictions. Exciting possibilities for molecular optics and a new type of optical chromatography are opened up.

Journal Article↗

PubMed2000

Preparation and biological activity of amino acid and peptide conjugates of antitumor hydroxymethylacylfulvene.

The primary hydroxyl group in hydroxymethylacylfulvene, a potent antitumor drug, is readily replaced by thiols including cysteine, N-acetylcysteine, homocysteine, and glutathione. Best yields are obtained when reaction is carried out in the presence of dilute sulfuric acid. A variety of sulfur-containing analogues have been prepared, and their toxicity to tumor cells was examined.

Amino Acids↗

PubMed2000

Cloning of a sugar utilization gene cluster in Aspergillus parasiticus.

At one end of the 70 kb aflatoxin biosynthetic pathway gene cluster in Aspergillus parasiticus and Aspergillus flavus reported earlier, we have cloned a group of four genes that constitute a well-defined gene cluster related to sugar utilization in A. parasiticus: (1) sugR, (2) hxtA, (3) glcA and (4) nadA. No similar well-defined sugar gene cluster has been reported so far in any other related Aspergillus species such as A. flavus, A. nidulans, A. sojae, A. niger, A. oryzae and A. fumigatus. The expression of the hxtA gene, encoding a hexose transporter protein, was found to be concurrent with the aflatoxin pathway cluster genes, in aflatoxin-conducive medium. This is significant since a close linkage between the two gene clusters could potentially explain the induction of aflatoxin biosynthesis by simple sugars such as glucose or sucrose.

Aflatoxins↗

PubMed2000

Does the completeness of prostate sampling predict outcome for patients undergoing radical prostatectomy?: data from the CAPSURE database.

OBJECTIVES: To determine whether more complete sampling of the radical prostatectomy (RP) specimen better predicts outcome after surgery. METHODS: We reviewed pathology reports from 1383 patients enrolled in CaPSURE (a longitudinal registry of patients with prostate cancer) who underwent RP. Specimens were considered step-sectioned only if the entire specimen was submitted for analysis and if sections were taken at 0.5-cm intervals or less. Otherwise, specimens were considered non-step-sectioned. Pathologic stage, Gleason score, surgical margin status, and outcome were compared between groups. Prostate-specific antigen (PSA) recurrence was defined as a PSA level of 0.2 ng/mL or greater on two consecutive occasions after RP. Secondary cancer treatment consisted of radiation or androgen deprivation after RP. Adjuvant treatments occurred within 6 months of RP, and nonadjuvant treatments occurred more than 6 months after RP. Kaplan-Meier event rates of PSA recurrence and secondary treatment were calculated for patients in the step-sectioned and non-step-sectioned groups. RESULTS: No significant differences were found between patients in the step-sectioned and non-step-sectioned groups with respect to pathologic tumor stage, prostatectomy Gleason score, or margin status. Patients in whom step-sectioning was performed had a lower serum PSA at diagnosis than patients in the non-step-sectioned group. When examining all patients, no differences were observed in the use of secondary treatments or PSA recurrence based on the method of pathologic analysis. However, patients with negative margins in whom step-sectioning was performed exhibited significantly lower secondary nonadjuvant treatment use and appeared to have a lower risk of PSA recurrence than similar patients in the non-step-sectioned group. CONCLUSIONS: These data suggest that more complete pathologic analysis of the surgical specimen may better predict outcome for some patients undergoing RP. Additional research is warranted to determine whether such differences justify the additional resources necessary to recommend routine step-sectioning.

Adult↗

PubMed2000

Gene expression profiling: monitoring transcription and translation products using DNA microarrays and proteomics.

Novel and powerful technologies such as DNA microarrays and proteomics have made possible the analysis of the expression levels of multiple genes simultaneously both in health and disease. In combination, these technologies promise to revolutionize biology, in particular in the area of molecular medicine as they are expected to reveal gene regulation events involved in disease progression as well as to pinpoint potential targets for drug discovery and diagnostics. Here, we review the current status of these technologies and highlight some studies in which they have been applied in concert to the analysis of biopsy specimens.

Animals↗

PubMed2000

Unmasking electronic energy transfer of conjugated polymers by suppression of O(2) quenching

The photochemistry of poly[2-methoxy, 5-(2'-ethyl-hexyloxy)-p-phenylene-vinylene] (MEH-PPV) has been found to be highly dependent on the presence of O(2), which increases singlet exciton quenching dramatically. Spectroscopy on isolated single molecules of MEH-PPV in polycarbonate films that exclude O(2) reveals two distinct polymer conformations with fluorescence maxima near 555 and 580 nanometers wavelength, respectively. Time-resolved single-molecule data demonstrate that the 580-nanometer conformation exhibits a "landscape" for intramolecular electronic energy relaxation with a "funnel" that contains a 580-nanometer singlet exciton trap at the bottom. The exciton traps can be converted to exciton quenchers by reaction with O(2). Conformationally induced, directed-energy transfer is arguably a critical dynamical process that is responsible for many of the distinctive photophysical properties of conjugated polymers.

Journal Article↗

PubMed2000

p53-dependent expression of PIG3 during proliferation, genotoxic stress, and reversible growth arrest.

The p53-inducible gene 3 (PIG3) was recently identified in a screen for genes induced by p53 before the onset of apoptosis. PIG3 shares significant homology with oxidoreductases from several species. In this study, PIG3-specific antibodies were used to analyze cellular PIG3 protein levels under control and genotoxic stress conditions. PIG3 protein was localized to the cytoplasm and induced in primary, non-transformed, and transformed cell cultures after exposure to genotoxic agents. The induction of PIG3 was p53-dependent and occurred with delayed kinetics as compared with other p53 downstream targets, such as p21 and MDM2. Using a p53-inducible cell model system, in which p53-mediated growth arrest is reversible, we found that PIG3 levels were increased during p53-mediated growth arrest. Interestingly, elevated levels of PIG3 were maintained in cells that resumed cycling in the absence of ectopic p53 expression, suggesting that PIG3 is a long-lived reporter, which may be useful for detecting transient activation of p53.

Apoptosis↗

PubMed2000

Cloning and mapping of human PKIB and PKIG, and comparison of tissue expression patterns of three members of the protein kinase inhibitor family, including PKIA.

Two novel members of the human cAMP-dependent protein kinase inhibitor (PKI) gene family, PKIB and PKIG, were cloned. The deduced proteins showed 70% and 90% identity with mouse PKIbeta and PKIgamma respectively. Both the already identified pseudosubstrate site and leucine-rich nuclear export signal motifs were defined from the 11 PKIs of different species. The PKIB and PKIG genes were mapped respectively to chromosome 6q21-22.1, using a radiation hybrid GB4 panel, and to chromosome 20q13.12-13.13, using a Stanford G3 panel. Northern-blot analysis of three PKI isoforms, including the PKIA identified previously, revealed significant differences in their expression patterns. PKIB had two transcripts of 1.9 kb and 1.4 kb. The former transcript was abundant in both placenta and brain and the latter was expressed most abundantly in placenta, highly in brain, heart, liver, pancreas, moderately in kidney, skeletal muscle and colon, and very little in the other eight tissues tested. PKIG was widely expressed as a 1.5-kb transcript with the highest level in heart, hardly detectable in thymus and peripheral blood leucocytes and was moderately expressed in the other tissues, with slightly different levels. However, PKIA was specifically expressed as two transcripts of 3.3 kb and 1.5 kb in heart and skeletal muscle. The distinct expression patterns of the three PKIs suggest that their roles in various tissues are probably different.

Adaptor Proteins, Signal Transducing↗

PubMed2000

Enhanced backward-directed multiphoton-excited fluorescence from dielectric microcavities

We demonstrate theoretically and experimentally that one-, two-, and three-photon excited fluorescence from dye molecules in spherical microcavities has an asymmetrical angular distribution and is enhanced in the backward direction. The enhancement ratios (of intensities at 180 degrees and 90 degrees ) are 9, 5, and 1.8 for three-, two-, and one-photon excitation, respectively. Even larger ratios are expected for microspheres with an index of refraction larger than that used in the experiments. Because of the reciprocity principle and concentration of the incident wave inside particles, the backward enhancement is expected to occur even with nonspherical particles.

Journal Article↗

PubMed2000

Collapse of stiff conjugated polymers with chemical defects into ordered, cylindrical conformations

The optical, electronic and mechanical properties of synthetic and biological materials consisting of polymer chains depend sensitively on the conformation adopted by these chains. The range of conformations available to such systems has accordingly been of intense fundamental as well as practical interest, and distinct conformational classes have been predicted, depending on the stiffness of the polymer chains and the strength of attractive interactions between segments within a chain. For example, flexible polymers should adopt highly disordered conformations resembling either a random coil or, in the presence of strong intrachain attractions, a so-called 'molten globule'. Stiff polymers with strong intrachain interactions, in contrast, are expected to collapse into conformations with long-range order, in the shape of toroids or rod-like structures. Here we use computer simulations to show that the anisotropy distribution obtained from polarization spectroscopy measurements on individual poly[2-methoxy-5-(2'-ethylhexyl)oxy-1,4-phenylenevinylene] polymer molecules is consistent with this prototypical stiff conjugated polymer adopting a highly ordered, collapsed conformation that cannot be correlated with ideal toroid or rod structures. We find that the presence of so-called 'tetrahedral chemical defects', where conjugated carbon-carbon links are replaced by tetrahedral links, divides the polymer chain into structurally identifiable quasi-straight segments that allow the molecule to adopt cylindrical conformations. Indeed, highly ordered, cylindrical conformations may be a critical factor in dictating the extraordinary photophysical properties of conjugated polymers, including highly efficient intramolecular energy transfer and significant local optical anisotropy in thin films.

Journal Article↗

PubMed2000

Platelet-derived growth factor (PDGF) receptor-alpha activates c-Jun NH2-terminal kinase-1 and antagonizes PDGF receptor-beta -induced phenotypic transformation.

Platelet-derived growth factor (PDGF) is a potent mitogen for mesenchymal cells. The PDGF B-chain (c-sis proto-oncogene) homodimer (PDGF BB) and v-sis, its viral counterpart, activate both alpha- and beta-receptor subunits (alpha-PDGFR and beta-PDGFR) and mediate anchorage-independent growth in NIH3T3 cells. In contrast, the PDGF A chain homodimer (PDGF AA) activates alpha-PDGFR only and fails to induce phenotypic transformation. In the present study, we investigated alpha- and beta-PDGFR specific signaling pathways that are responsible for the differences between the transforming ability of PDGF AA and BB. To study PDGF BB activation of beta-PDGFR, we established NIH3T3 clones in which alpha-PDGFR signaling is inhibited by a dominant-negative alpha-PDGFR, or an antisense construct of alpha-PDGFR. Here, we demonstrate that beta-PDGFR activation alone is sufficient for PDGF BB-mediated anchorage-independent cell growth. More importantly, inhibition of alpha-PDGFR signaling enhanced PDGF BB-mediated phenotypic transformation, suggesting that alpha-PDGFR antagonizes beta-PDGFR-induced transformation. While both alpha- and beta-receptors effectively activate ERKs, alpha-PDGFR, but not beta-PDGFR, activates stress-activated protein kinase-1/c-Jun NH(2)-terminal kinase-1 (JNK-1). Inhibition of JNK-1 activity using a dominant-negative JNK-1 mutant markedly enhanced PDGF BB-mediated anchorage-independent cell growth, demonstrating an antagonistic role for JNK-1 in PDGF-induced transformation. Consistently, overexpression of wild-type JNK-1 reduced PDGF BB-mediated transformation. Taken together, the present study showed that alpha- and beta-PDGFRs differentially regulate Ras-mitogen-activated protein kinase pathways critical for regulation of cell transformation, and transformation suppressing activity of alpha-PDGFR involves JNK-1 activation.

3T3 Cells↗

PubMed2000

Effects of exercise training on left ventricular function and peripheral resistance in patients with chronic heart failure: A randomized trial.

CONTEXT: Exercise training in patients with chronic heart failure improves work capacity by enhancing endothelial function and skeletal muscle aerobic metabolism, but effects on central hemodynamic function are not well established. OBJECTIVE: To evaluate the effects of exercise training on left ventricular (LV) function and hemodynamic response to exercise in patients with stable chronic heart failure. DESIGN: Prospective randomized trial conducted in 1994-1999. SETTING: University department of cardiology/outpatient clinic in Germany. PATIENTS: Consecutive sample of 73 men aged 70 years or younger with chronic heart failure (with LV ejection fraction of approximately 0.27). INTERVENTION: Patients were randomly assigned to 2 weeks of in-hospital ergometer exercise for 10 minutes 4 to 6 times per day, followed by 6 months of home-based ergometer exercise training for 20 minutes per day at 70% of peak oxygen uptake (n=36) or to no intervention (control group; n=37). MAIN OUTCOME MEASURES: Ergospirometry with measurement of central hemodynamics by thermodilution at rest and during exercise; echocardiographic determination of LV diameters and volumes, at baseline and 6-month follow-up, for the exercise training vs control groups. RESULTS: After 6 months, patients in the exercise training group had statistically significant improvements compared with controls in New York Heart Association functional class, maximal ventilation, exercise time, and exercise capacity as well as decreased resting heart rate and increased stroke volume at rest. In the exercise training group, an increase from baseline to 6-month follow-up was observed in mean (SD) resting LV ejection fraction (0.30 [0.08] vs 0.35 [0.09]; P=.003). Mean (SD) total peripheral resistance (TPR) during peak exercise was reduced by 157 (306) dyne/s/cm(-5) in the exercise training group vs an increase of 43 (148) dyne/s/cm(-5) in the control group (P=.003), with a concomitant increase in mean (SD) stroke volume of 14 (22) mL vs 1 (19) mL in the control group (P=.03). There was a small but significant reduction in mean (SD) LV end diastolic diameter of 4 (6) mm vs an increase of 1 (4) mm in the control group (P<.001). Changes from baseline in resting TPR for both groups were correlated with changes in stroke volume (r=-0.76; P<.001) and in LV end diastolic diameter (r=0.45; P<.001). CONCLUSIONS: In patients with stable chronic heart failure, exercise training is associated with reduction of peripheral resistance and results in small but significant improvements in stroke volume and reduction in cardiomegaly. JAMA. 2000.

Aged↗

PubMed2000

Cloning and characterization of avfA and omtB genes involved in aflatoxin biosynthesis in three Aspergillus species.

The biosynthesis of aflatoxins (B(1), G(1), B(2), and G(2)) is a multi-enzyme process controlled genetically by over 20 genes. In this study, we report the identification and characterization of the avfA gene, which was found to be involved in the conversion of averufin (AVF) to versiconal hemiacetal acetate (VHA), in Aspergillus parasiticus and A. flavus; a copy of avfA gene was also cloned from a non-aflatoxin producing strain A. sojae. Complementation of an averufin-accumulating, non-aflatoxigenic mutant strain of A. parasiticus, SRRC 165, with the avfA gene cloned from A. flavus, restored the ability of the mutant to convert AVF to VHA and to produce aflatoxins B(1), G(1), B(2), and G(2). Sequence analysis revealed that a single amino acid replacement from aspartic acid to asparagine disabled the function of the enzyme in the mutant strain SRRC 165. The A. parasiticus avfA was identified to be a homolog of previously sequenced, but functionally unassigned transcript, stcO, in A. nidulans based on sequence homology at both nucleotide (57%) and amino acid (55%) levels. In addition to avfA, another aflatoxin pathway gene, omtB, encoding for an O-methyltransferase involved in the conversion of demethylsterigmatocystin (DMST) to sterigmatocystin (ST) and dihydrodemethylsterigmatocystin (DHDMST) to dihydrosterigmatocystin (DHST), was cloned from A. parasiticus, A. flavus, and A. sojae. The omtB gene was found to be highly homologous to stcP from A. nidulans, which has been reported earlier to be involved in a similar enzymatic step for the sterigmatocystin formation in that species. RT-PCR data demonstrated that both the avfA and avfA1 as well as omtB genes in A. parasiticus were expressed only in the aflatoxin-conducive medium. An analysis of the degrees of homology for the two reported genes between the Aspergillus species A. parasiticus, A. flavus, A. nidulans and A. sojae was conducted.

Aflatoxins↗

PubMed2000

Characterization of the Aspergillus parasiticus major nitrogen regulatory gene, areA.

The major nitrogen regulatory gene, areA, was cloned from Aspergillus parasiticus. It encoded a polypeptide of 864 amino acids which contained a nuclear localization signal (NLS), a highly acidic region from positions 497 to 542, a Cys-X(2)-Cys-X(17)-Cys-X(2)-Cys DNA-binding motif and a conserved carboxy-terminus. Electrophoretic mobility shift assays suggested that the A. parasiticus AREA DNA-binding domain fusion protein bound cooperatively to single GATA elements in the A. parasiticus niaD-niiA intergenic region. AREA also bound to the aflR-aflJ intergenic region of the aflatoxin biosynthesis gene cluster. Regions of areA were fused to a yeast GAL4 DNA-binding domain coding region to localize putative transcription activation domain(s) of AREA based on activation of the GAL1(p)::lacZ reporter gene expression. The portion between NLS and the acidic domain demonstrated 16-20-fold higher activation activities than other portions of AREA, which suggests that the transcription activation domain is located in this region.

Aspergillus↗

PubMed2000

Molecular separation of two behavioral phenotypes by a mutation affecting the promoters of a Ca-activated K channel.

The Drosophila slowpoke gene encodes a BK-type calcium-activated potassium channel. Null mutations in slowpoke perturb the signaling properties of neurons and muscles and cause behavioral defects. The animals fly very poorly compared with wild-type strains and, after exposure to a bright but cool light or a heat pulse, exhibit a "sticky-feet" phenotype. Expression of slowpoke arises from five transcriptional promoters that express the gene in neural, muscle, and epithelial tissues. A chromosomal deletion (ash2(18)) has been identified that removes the neuronal promoters but not the muscle-tracheal cell promoter. This deletion complements the flight defect of slowpoke null mutants but not the sticky-feet phenotype. Electrophysiological assays confirm that the ash2(18) chromosome restores normal electrical properties to the flight muscle. This suggests that the flight defect arises from a lack of slowpoke expression in muscle, whereas the sticky-feet phenotype arises from a lack of expression in nervous tissue.

Action Potentials↗