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B Zhao

Publications and source records attributed to B Zhao.

At least 73 records · Page 4Linked to original sources
PubMed2000

Stabilization of bound polycyclic aromatic hydrocarbons by a pi-cation interaction.

Proteins can use aromatic side-chains to stabilize bound cationic ligands through cation-pi interactions. Here, we report the first example of the reciprocal process, termed pi-cation, in which a cationic protein side-chain stabilizes a neutral aromatic ligand. Site-directed mutagenesis revealed that an arginine side-chain located in the deep binding pocket of a monoclonal antibody (4D5) is essential for binding the neutral polynuclear aromatic hydrocarbon benzo[a]pyrene. This Arg was very likely selected for in the primary response, further underscoring the importance of the pi-cation interaction for ligand binding, which should be considered in protein analysis and design when ligands include aromatic groups.

Amino Acid Sequence↗
PubMed2000

The statistical analysis of the in vitro chromosome aberration assay using Chinese hamster ovary cells.

The purpose of the in vitro chromosome aberration assay (ABS) is to determine whether the test compound is a clastogen, i.e. induces structural changes in chromosomes. Details of this assay can be found in Galloway et al. [S.M. Galloway, M. Aardema, M. Ishidate Jr, J.L. Ivett, D.J. Kirkland, M. Takeshi, P. Mosesso, T. Sofuni, Mutation Res. 312 (1994) 241-261]. The standard design consists of a negative control and at least three positive dose groups. At each dose, a sample, say 200, of metaphase cells is examined microscopically and cells exhibiting at least one type of chromosome aberration are identified. Using Chinese hamster ovary cells, Margolin et al. [B.H. Margolin, M.A. Resnick, J.Y. Rimpo, P. Archer, S.M. Galloway, A.D. Bloom, E. Zeiger, Environ. Mutagen. 8 (1986) 183-204] and Richardson et al. [C. Richardson, D.A. Williams, J.A. Allen, G. Amphlett, D.O. Chanter, B. Phillips, Analysis of data from in vitro cytogenetic assays, in: D.J. Kirkland (Ed.), Statistical Evaluation of Mutagenicity Test Data, Cambridge University Press, Cambridge, 1989, pp. 141-154] demonstrated that a binomial sampling model could be used to describe the proportion of cells with chromosome aberrations.Statisticians and toxicologists have also suggested evaluation criteria for the dose response pattern of ABS. Margolin et al. [B.H. Margolin, M.A. Resnick, J.Y. Rimpo, P. Archer, S.M. Galloway, A.D. Bloom, E. Zeiger, Environ. Mutagen. 8 (1986) 183-204] suggested one use the Cochran-Armitage trend test. Sofuni et al. [T. Sofuni, A. Matsuoka, M. Sawada, M. Ishidate Jr, E. Zeiger, M.D. Shelby, Mutation Res. 241 (1990) 175-213] considered the dose response to be (strong) positive if it had two significant doses out of three dose groups and decided it was weakly positive if it had only one significant dose and there was a significant trend. The criterion of Galloway et al. for a positive response was a clear dose-related increase in cells with structural aberrations in one experiment or a reproducible single positive dose [S.M. Galloway, M. Aardema, M. Ishidate Jr, J.L. Ivett, D.J. Kirkland, M. Takeshi, P. Mosesso, T. Sofuni, Mutation Res. 312 (1994) 241-261]. We formulate the above three procedures in terms of a Cochran-Armitage trend test and a Dunnett type test. We then compare the performance of these three procedures in terms of a Monte Carlo simulation study. We then develop a software program from the chosen procedure for its ease of use by statisticians and toxicologists.

Animals↗
PubMed2000

Modular bacterial artificial chromosome vectors for transfer of large inserts into mammalian cells.

To facilitate the use of large-insert bacterial clones for functional analysis, we have constructed new bacterial artificial chromosome vectors, pPAC4 and pBACe4. These vectors contain two genetic elements that enable stable maintenance of the clones in mammalian cells: (1) The Epstein-Barr virus replicon, oriP, is included to ensure stable episomal propagation of the large insert clones upon transfection into mammalian cells. (2) The blasticidin deaminase gene is placed in a eukaryotic expression cassette to enable selection for the desired mammalian clones by using the nucleoside antibiotic blasticidin. Sequences important to select for loxP-specific genome targeting in mammalian chromosomes are also present. In addition, we demonstrate that the attTn7 sequence present on the vectors permits specific addition of selected features to the library clones. Unique sites have also been included in the vector to enable linearization of the large-insert clones, e. g., for optical mapping studies. The pPAC4 vector has been used to generate libraries from the human, mouse, and rat genomes. We believe that clones from these libraries would serve as an important reagent in functional experiments, including the identification or validation of candidate disease genes, by transferring a particular clone containing the relevant wildtype gene into mutant cells or transgenic or knock-out animals.

Animals↗
PubMed2000

Mechanisms of apoptosis in rat cerebellar granule cells induced by hydroxyl radicals and the effects of EGb761 and its constituents.

In this study investigation is made on whether oxidative stress produced by treatment with hydroxyl radicals can induce apoptosis in rat cerebellar granule cells. The protective effects of Ginkgo biloba extract (EGb761) and its active constituents against apoptosis are also examined. The results show that hydroxyl radicals generated by the Fenton reaction induced apoptosis in cerebellar granule cells, which was associated with the decrease in the Bcl-2 mRNA level and the increase in the protein levels of the transcription factors Fos and Jun. Moreover, hydroxyl radicals induced time-dependent lipid peroxidation in cells and caused the changes in the sulfhydryl group binding sites on the membrane proteins. Hydroxyl radicals may induce apoptosis via different signaling pathways. EGb761 attenuated these changes and its different constituents showed different effects. The total flavonoid component of EGb761 and a mixture of flavonoids and terpenes protected cerebellar granule cells from oxidative damage and apoptosis induced by hydroxyl radicals. Total terpenes of EGb761 did not protect against apoptosis. Flavonoids and terpenes did not show a synergistic effect in this regard.

Animals↗
PubMed2000

Constitutive achaete-scute homologue-1 promotes airway dysplasia and lung neuroendocrine tumors in transgenic mice.

The transcription factor achaete-scute homologue-1 (ASH1) is essential for neural differentiation during fetal development and is a cardinal feature of neuroendocrine (NE) tumors such as small cell lung cancer. To explore the potential of ASH1 to promote NE differentiation and tumorigenesis in the lung, we constitutively expressed the factor in nonendocrine airway epithelial cells using transgenic mice. Progressive airway hyperplasia and metaplasia developed beginning at 3 weeks of life. ASH1 potently enhanced the tumorigenic effect of SV40 large T antigen in airway epithelium. These doubly transgenic animals developed massive NE lung tumors, implying that ASH1 may cooperate with defects in p53, pRb, or related pathways in promoting NE lung carcinogenesis.

Animals↗
PubMed2000

A negative regulatory element within the proximal promoter region of the rat ornithine decarboxylase gene.

A putative Ets site with a core of GGAA located at nt -88 to -85 of the rat ornithine decarboxylase (ODC) gene was characterized by site-directed mutagenesis and transient expression assays. Mutation of this site, when in pODClux2m, which contains a cluster of four Sp1-binding sites, resulted in a 2.6-fold increase in basal promoter activity in untreated cells, whereas the ratio of activity in 12-O-tetradecanoylphorbol-13-acetate (TPA)-treated cells relative to the ratio in untreated cells (the induction ratio) remained largely unchanged. However, when the mutation was in pODClux168, which contains only a single Sp1-binding site (GC box V), it caused little alteration to either basal promoter activity or TPA induction ratio. A protein of 55-60 kDa was found specifically bound to this site, as shown by ultraviolet cross-linking assay. In competition assay and methylation interference assay, this protein was shown to occupy the GGAA core, although it showed no antigenic relation to c-Ets-1 in an supershift assay. We suggest that this protein binds specifically to the GGAA core and functions to inhibit activation of the ODC promoter by distal elements, including the upstream Sp1 sites.

Animals↗
PubMed2000

Hydrogen peroxide-induced oxidative damage and apoptosis in cerebellar granule cells: protection by Ginkgo biloba extract.

The ability of oxidative stress to induce apoptosis and the protective effects of Ginkgo biloba extract (EGb761) against this induction were studied in cultures of rat cerebellar granule cells. Cells were exposed to oxidative stress by treatment with 50 microm hydrogen peroxide+100 microm ferrous sulphate which generates hydroxyl radicals by Fenton reaction. Both morphological observation and biochemical analysis revealed that H(2)O(2)/FeSO(4)treatment induced apoptotic cell death in cerebellar granule cells, which was characterized by chromatin condensation and DNA fragmentation. During this process, the fluidity of the cell membrane decreased markedly, and the conformation of membrane proteins altered significantly. Pretreating cerebellar granule cells with the antioxidant EGb761 (Ginkgo biloba extract) effectively attenuated oxidative damage induced by H(2)O(2)/FeSO(4), and prevented cells from apoptotic cell death. The results suggested that EGb761 might be used as a potential drug for neuronal diseases associated with the excessive production of reactive oxygen species.

Animals↗
PubMed2000

Correlation between acetylation phenotype and genotype in Chinese women.

OBJECTIVE: The acetylation polymorphism is a common inherited variation in human drug and carcinogen metabolism. Because N-acetyltransferase (NAT2) is important for the detoxification and/or bioactivation of drugs and carcinogens, this polymorphism has important implications in therapeutics and cancer susceptibility. A high correlation between acetylation phenotype and genotype has been demonstrated in several studies. However, no such data exist for Chinese females. The aim of the present study was to compare acetylation phenotype with NAT2 genotype in a population of primarily non-smoking Chinese females. METHODS: In the present study, the correlation between N-acetyltransferase activity and NAT2 genotype was evaluated in 103 unrelated Chinese female controls derived from a hospital-based case-control study of lung cancer in Singapore. Acetylation phenotype and genotype were respectively determined using caffeine and an allele-specific polymerase chain reaction (PCR). RESULTS: The proportions of rapid and slow phenotypes were 78% and 22%, respectively, while the distribution of rapid (heterozygotes and homozygotes combined) and slow acetylator genotypes was 76% and 24%, respectively. The distribution of the various NAT2 genotypes did not differ significantly (chi2 = 1.45, P > 0.05) from that predicted by the Hardy-Weinberg Law. All slow acetylators were accurately predicted (100%), whereas 2 of 80 rapid acetylators were erroneously predicted as slow (2.5%). The overall prediction rate of the PCR-based test for the acetylation phenotype was at 98.1% in our Chinese population. CONCLUSION: Our results suggest that genotyping with PCR may well become the preferred method for the determination of acetylation polymorphism in epidemiological studies in this Asian population.

Acetylation↗
PubMed2000

Significance of K-ras mutation and CEA level in pancreatic juice in the diagnosis of pancreatic cancer.

The early diagnosis of pancreatic carcinoma is essential for increasing patient survival rates. In this study, 52 patients with suspected pancreatic diseases were examined to investigate the value of K-ras codon 12 point mutation, levels of carcinoembryonic antigen (CEA) and carbohydrate antigen (CA19-9), and cytology of pancreatic juice in the diagnosis of pancreatic carcinoma. Pancreatic juice was taken without secretin stimulation. K-ras mutation was detected by enriched polymerase chain reaction (PCR) restriction fragment length polymorphism (RFLP). K-ras mutation in pancreatic juice was more frequent in carcinoma than in benign diseases (P = 0.0448). The positive predictive value of K-ras mutation for the diagnosis of neoplastic disease was 83%. The CEA level in pancreatic juice in carcinoma was significantly greater than that in benign disease (P< 0.0001). When the cutoff level of CEA was set at 50 ng/ml, its accuracy for the diagnosis of carcinoma was 85%. A multivariate analysis showed that K-ras mutation and CEA level in pancreatic juice, as well as serum CA19-9 level and age of the patient were independent variables for the diagnosis of carcinoma, and the accuracy of diagnosis by this analysis was increased to 90%. In conclusion, both K-ras mutation and CEA level in pancreatic juice may be valuable for the diagnosis of carcinoma. Better discrimination was possible with a multivariate analysis.

Adult↗
PubMed2000

Ab initio studies on the vibrational spectra and conformational isomerism of 2,4-dimethyl-1,3-pentadiene.

Ab initio was used to study the structure of various conformational isomers and their vibrational spectra of 2,4-dimethyl-1,3-pentadiene (2,4-PD) in detail. Two stable conformations, s-trans and s-cis, were found in which s-trans is more stable. The geometry of stable conformations and charge distributions were studied, and the effect of different basis sets on geometry optimization is discussed. The results of complete optimization indicate that molecular skeleton is nearly in a plane, its largest deviation is only 0.3 degrees. Therefore, it is reasonable and available to hypothesis that the molecule has Cs symmetry. The thermal dynamics conformations were calculated and compared with experimental values. DeltaH(o) between two conformations of 2,4-PD measured from experiment is 4.36 kJ/mol, deltaS(o) is 2.56 J/mol K., calculated results are slightly different from experimental ones. Vibrational frequencies of 2,4-PD conformers have been studied by ab initio molecular orbital calculations using different basis set. The calculated vibrational frequencies are analyzed and compared with the experimental spectra.

Alkadienes↗
PubMed2000

Two novel secoergosterols from the fungus Tylopilus plumbeoviolaceus.

Two novel secoergosterols, 3beta-hydroxy-8alpha,9alpha-oxido-8, 9-secoergosta-7,9(11),22-triene (tylopiol A) (1) and 3beta-hydroxy-8alpha,9alpha-oxido-8,9-secoergosta-7,22 -dien-12-one (tylopiol B) (2), were isolated from the fresh fruit bodies of Tylopilus plumbeoviolaceus, along with three known compounds, ergosta-7,22-dien-3beta-ol, uridine, and allitol. Their structures were elucidated by NMR techniques, including (1)H NMR, (13)C NMR, HMQC, HMBC, and MS. The structure and stereochemistry of compound 1 were demonstrated by X-ray crystallography.

Agaricales↗
PubMed2000

Lipoprotein lipase-mediated interactions of small proteoglycans and low-density lipoproteins.

According to numerous studies low-density lipoproteins (LDL) are supposed to interact with the glycosaminoglycan chain(s) of proteoglycans, e.g. with decorin and biglycan, which themselves are subject to receptor-mediated endocytosis. We tested, therefore, whether complexes of LDL and small proteoglycans can be endocytosed by either the LDL- or the small proteoglycan uptake mechanism. However, neither was the endocytosis of LDL significantly influenced by proteoglycans nor that of proteoglycans by LDL. This negative result could be explained by the observation that in vitro complex formation takes place only in buffers of low ionic strength. Under physiological conditions additional molecules may be necessary for complex stabilization. Lipoprotein lipase (LpL) which binds LDL was also able to interact with high affinity with decorin and its glycosaminoglycan-free core protein, both interactions being heparin-sensitive. Regardless of the presence or absence of LDL, LpL stimulated the endocytosis of decorin 1.5-fold, whereas LpL mediated a 4-fold stimulation of LDL uptake in the absence of decorin. No significant additional effect was seen in the presence of small concentrations of proteoglycans whereas in the presence of 1 microM decorin the endocytosis of [125I]LDL was reduced in normal as well as in LDL receptor-deficient fibroblasts. These observations could best be explained by assuming that LpL/LDL complexes are internalized upon binding to membrane-associated heparan sulphate and that small proteoglycans interfere with this process. It could not be ruled out, however, that a small proportion of the complexes is also taken up by the small proteoglycan receptor.

Biglycan↗
PubMed2000

Transgenic mouse models for lung cancer.

Transgenic technology allows the ability to target regulatory genes to the lungs in a cell-specific fashion. Using this technology, we have generated a model to investigate the phenotypic consequences of targeting oncogenes to particular cell types in the lungs and are developing a second model for the regulated expression of oncogenes in the lung. The transgenic model involves the constitutive expression of simian virus 40 large T antigen in the Clara cells of mouse lungs. This model has been used to investigate changes in expression of cell cycle regulatory genes in the Clara cells during the transformation process, as well as the expression of the transcription factors regulating the expression of Clara cell differentiation markers. The second model we are developing investigates the regulated expression of the genes in the lungs. This system is based on the establishment of two types of transgenic lines. The regulator line consists of a chimeric transcriptional factor placed under the control of a lung-specific SPC (surfactant protein C) promoter. This chimeric regulator is composed of a transcription activation domain, the GAL4 DNA-binding domain, and a truncated progesterone receptor that is responsive to RU 486, but not to endogenous progesterone. The second transgenic mouse line contains the silent target transgene under the control of a minimal promoter with upstream activating sequences (UAS) that are recognized by the regulator transgene. Upon breeding these two lines, the resulting bitransgenic mice can then be induced to express the target transgene only with the administration of RU 486. Two generations of regulators have been evaluated on their ability to regulate the expression of a growth hormone reporter gene. This system demonstrated the inducible expression of the reporter genes in the distal airways of the lungs.

Adenocarcinoma↗
PubMed2000

Significance of thymidine phosphorylase/platelet-derived endothelial cell growth factor in carcinoma of the papilla of Vater.

The expression of thymidine phosphorylase (TP) in carcinoma of the papilla of Vater was studied to clarify its significance in tumor progression and in determining prognosis. Fifty-nine cases of surgically resected carcinoma of the papilla of Vater were studied. Immunohistochemical staining was performed to evaluate the expression of TP, microvessel count and p53 overexpression. TP expression was demonstrated in tumor cells in 62.7% (37/59) of the cases. A higher frequency of regional lymph node metastasis was found in TP-positive tumors than in TP-negative tumors (P = 0.006). TP-positive tumors were more advanced than TP-negative tumors with regard to clinical stage (P = 0.035). TP-positive tumors had significantly higher microvessel density (27.6 +/- 10.1) than TP-negative tumors (20.4 +/- 10.0, P = 0.01). Moreover, TP expression was significantly correlated with a poor prognosis (P = 0. 02). These suggest that in carcinoma of the papilla of Vater, TP production by tumor cells is correlated with tumor progression through its regulatory effect on neovascularization.

Adult↗
PubMed2000

Physiological regulation of uteroglobin/CCSP expression.

Uteroglobin/CCSP is expressed specifically in the Clara cells. This allows the gene to be used as a marker to identify the elements regulating the physiologic and cell-specific expression of this gene. The regulation of UG/CCSP by IFN-gamma was shown to be at the level of the proximal promoter by the upregulation of HNF3 beta. This has allowed the determination of the factors responsible for the expression of UG/CCSP.

Animals↗