Search PubMed⌕ Search

Biomedical subjects

Wei Yang

Publications and source records attributed to Wei Yang.

At least 109 records · Page 6Linked to original sources
PubMed2005

Probing site-specific calmodulin calcium and lanthanide affinity by grafting.

Ca2+ binding is essential for the biological functions of calmodulin (CaM) as a trigger/sensor protein to regulate many biological processes in the Ca2+ -signaling cascade. A challenge in understanding the mechanism of Ca2+ signaling is to obtain site-specific information about the Ca2+ binding properties of individual Ca2+ -binding sites of EF-hand proteins, especially for CaM. In this paper, we report the first estimation of the intrinsic Ca2+ affinities of the four EF-hand loops of calmoduin (I-IV) by individually grafting into the domain 1 of CD2. Taking advantage of the Trp residues in the host protein, we first determined metal-binding affinities for Tb3+, Ca2+, and La3+ for all four grafted EF-loops using Tb3+ aromatic resonance energy transfer. EF-loop I exhibits the strongest binding affinity for Ca2+, La3+, and Tb3+, while EF-loop IV has the weakest metal-binding affinity. EF-loops I-IV of CaM have dissociation constants for Ca2+ of 34, 245, 185, and 814 microM, respectively, with the order I > III approximately equal to II > IV. These findings support a charge-ligand-balanced model in which both the number of negatively charged ligand residues and the balanced electrostatic dentate-dentate repulsion by the adjacent charged residues are two major determinants for the relative Ca2+ -binding affinities of EF-loops in CaM. Our grafting method provides a new strategy to obtain site-specific Ca2+ binding properties and a better estimation of the cooperativity and conformational change contributions of coupled EF-hand proteins.

Animals↗
PubMed2005

Design of a calcium-binding protein with desired structure in a cell adhesion molecule.

Ca2+, "a signal of life and death", controls numerous cellular processes through interactions with proteins. An effective approach to understanding the role of Ca2+ is the design of a Ca2+-binding protein with predicted structural and functional properties. To design de novo Ca2+-binding sites in proteins is challenging due to the high coordination numbers and the incorporation of charged ligand residues, in addition to Ca2+-induced conformational change. Here, we demonstrate the successful design of a Ca2+-binding site in the non-Ca2+-binding cell adhesion protein CD2. This designed protein, Ca.CD2, exhibits selectivity for Ca2+ versus other di- and monovalent cations. In addition, La3+ (Kd 5.0 microM) and Tb3+ (Kd 6.6 microM) bind to the designed protein somewhat more tightly than does Ca2+ (Kd 1.4 mM). More interestingly, Ca.CD2 retains the native ability to associate with the natural target molecule. The solution structure reveals that Ca.CD2 binds Ca2+ at the intended site with the designed arrangement, which validates our general strategy for designing de novo Ca2+-binding proteins. The structural information also provides a close view of structural determinants that are necessary for a functional protein to accommodate the metal-binding site. This first success in designing Ca2+-binding proteins with desired structural and functional properties opens a new avenue in unveiling key determinants to Ca2+ binding, the mechanism of Ca2+ signaling, and Ca2+-dependent cell adhesion, while avoiding the complexities of the global conformational changes and cooperativity in natural Ca2+-binding proteins. It also represents a major achievement toward designing functional proteins controlled by Ca2+ binding.

Animals↗
PubMed2005

Anti-tumor effect of pEgr-interferon-gamma-endostatin gene-radiotherapy in mice bearing Lewis lung carcinoma and its mechanism.

BACKGROUND: Gene-radiotherapy, the combination of gene therapy and radiation therapy, is a new paradigm for cancer treatment. To enhance anti-tumor effect of gene-radiotherapy, in this study we construct a radiation-inducible dual-gene co-expression vector pEgr-interferon (IFN)-gamma-endostatin and studied the anti-tumor effect of pEgr-IFN-gamma-endostatin gene-radiotherapy in mice bearing Lewis lung carcinoma and its mechanism. METHODS: Gene recombinant technique was used to construct dual-gene co-expression plasmid pEgr-IFN-gamma-endostatin, and single-gene expression plasmid pEgr-IFN-gamma and pEgr-endostatin. The plasmids packed by liposome were injected locally into the tumors of the mice, and the tumors were irradiated with 5 Gy X-ray 36 hours later. The tumor growth rate at different time and mean survival period of the mice were observed. Cytotoxic activity of splenic cytotoxic T-lymphocyte (CTL), natural killer (NK) cell and tumor necrosis factor (TNF)-alpha secretion activity of peritoneal macrophages of the mice in various groups were evaluated 15 days after irradiation. The intratumor micro-vessel density was evaluated by immunohistochemical staining 10 days after irradiation. RESULTS: The tumor growth rate of the mice in dual-gene-radiotherapy group was significantly lower than those in control group, 5 Gy group and single-gene-radiotherapy group at different time after gene-radiotherapy, and the mean survival period of which was longer. Cytotoxic activity of splenic CTL, NK and TNF-alpha secretion activity of peritoneal macrophages of the mice in dual-gene-radiotherapy group were significantly higher than those in control group, 5 Gy X-ray irradiation group and pEgr-endostatin gene-radiotherapy group 15 days after irradiation. The intratumor micro-vessel density of the mice in dual-gene-radiotherapy group was significantly lower than those in control group, 5 Gy X-ray irradiation group and pEgr-IFN-gammagene-radiotherapy group. CONCLUSION: The anti-tumor effect of dual-gene-radiotherapy was significantly better than that of single-gene-radiotherapy by combining the enhancement of anti-tumor immunologic function induced by IFN-gamma with the anti-angiogenesis function of endostatin.

Animals↗
PubMed2005

Protein structural transitions and their functional role.

Living cells are a collection of molecular machines which carry out many of the functions essential for the cell's existence, differentiation and reproduction. Most, though not all, of these machines are made up of proteins. Because of their complexity, an understanding of how they work requires a synergistic combination of experimental and theoretical studies. In this paper we outline our studies of two such protein machines. One is GroEL, the chaperone from Escherichia coli, which aids in protein folding; the other is F(1)-ATPase, a motor protein which synthesizes and hydrolyses ATP.

Chaperonin 60↗
PubMed2005

Gene transfer to plants by diverse species of bacteria.

Agrobacterium is widely considered to be the only bacterial genus capable of transferring genes to plants. When suitably modified, Agrobacterium has become the most effective vector for gene transfer in plant biotechnology. However, the complexity of the patent landscape has created both real and perceived obstacles to the effective use of this technology for agricultural improvements by many public and private organizations worldwide. Here we show that several species of bacteria outside the Agrobacterium genus can be modified to mediate gene transfer to a number of diverse plants. These plant-associated symbiotic bacteria were made competent for gene transfer by acquisition of both a disarmed Ti plasmid and a suitable binary vector. This alternative to Agrobacterium-mediated technology for crop improvement, in addition to affording a versatile 'open source' platform for plant biotechnology, may lead to new uses of natural bacteria-plant interactions to achieve plant transformation.

Arabidopsis↗
PubMed2005

Portraits of a Y-family DNA polymerase.

Members of the Y-family of DNA polymerases catalyze template-dependent DNA synthesis but share no sequence homology with other known DNA polymerases. Y-family polymerases exhibit high error rates and low processivity when copying normal DNA but are able to synthesize DNA opposite damaged templates. In the past three years, much has been learned about this family of polymerases including determination of more than a dozen crystal structures with various substrates. In this short review, I will summarize the biochemical properties and structural features of Y-family DNA polymerases.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗
PubMed2005

Identification of gene expression modifications in myostatin-stimulated myoblasts.

Myostatin belongs to the transforming growth factor beta superfamily and has been shown to function as an inhibitor of skeletal muscle proliferation and differentiation. To gain insight into the molecular mechanisms of myostatin function during myogenesis, differential display reverse transcription PCR was employed to identify altered gene expressions associated with myostatin inhibitory function in chicken fetal myoblasts (CFMs). In this work, we have identified seven up-regulated and 12 down-regulated genes in myostatin stimulated CFMs. Those genes are involved in myogenic differentiation, cell architecture, energy metabolism, signal transduction, and apoptosis. The down-regulation of muscle creatine kinase B, troponin C, and myosin regulatory light chain is in agreement with the myostatin negative role in myocyte differentiation. In addition, the expression alteration of skeletal muscle-specific cardiac ankyrin repeat protein and the bcl-2 related anti-apoptotic protein Nr-13 suggests possible unique roles for myostatin in regulating myogenesis by controlling cofactors participated transcriptional regulation and apoptosis.

Animals↗
PubMed2005

Role of ERK1/2 in FSH induced PCNA expression and steroidogenesis in granulosa cells.

Follicular development is characterized by both proliferation and differentiation of granulosa cells (GCs) under the control of FSH. However, the cellular mechanism by FSH is not known. Using cultured GCs, we examined whether FSH activated ERK1/2 was involved in the regulation of the proliferation related gene proliferating cell nuclear antigen (PCNA) and steroidogenesis. GCs were obtained from the ovaries of DES treated immature rats and cultured in serum free medium. The results showed that FSH activated ERK1/2 in a time dependent manner, with a peak at 20 min. Such activation was PKA dependent as was inhibited by specific inhibitors. FSH induced PCNA expression in a time dependent manner, with a maximum stimulation at 2 h. Similarly, StAR and steroid levels increased as FSH treatment time extended, with a maximum progesterone and StAR production at 48 h. ERK1/2 inactivation by UO126 inhibited the stimulatory effects of FSH on both PCNA and StAR expression and steroid synthesis in the GCs (p less than 0.01). Immunocytochemical studies further revealed that ERK1/2 inhibition led to a reduction of mitochondrial StAR in the GCs by FSH. These observations suggested that the stimulation of FSH on PCNA expression and steroidogenesis in GCs was mediated at least partially by ERK1/2.

Animals↗
PubMed2005

Periconceptional dietary intake of myo-inositol and neural tube defects in offspring.

BACKGROUND: Periconceptional intake of nutrients in addition to folic acid may contribute to neural tube defect (NTD) etiologies; a likely candidate is myo-inositol. We investigated whether maternal periconceptional dietary intake of myo-inositol influenced NTD risk. METHODS: Data were derived from a case-control study of fetuses and infants with NTDs among 1989-1991 California births. Interviews were conducted with mothers of 454 NTD cases and with mothers of 462 nonmalformed controls. A standard 100-item food frequency questionnaire was used to assess nutrient intake. RESULTS: We observed small increases in risk, with increases slightly more evident for anencephaly, associated with intakes of myo-inositol less than the highest intake quartile, e.g., risk of anencephaly was 1.3 (0.7-2.4) among fetuses whose mothers consumed lowest versus highest intakes of myo-inositol. These small increases, however, were imprecise, and also did not indicate increasing risk with decreasing level of myo-inositol intake. Adjusted risk estimates did not differ considerably from their unadjusted counterparts. CONCLUSIONS: Our results do not indicate that myo-inositol intake, as measured in this study, is strongly associated with risk of human NTDs.

Female↗
PubMed2005

Diminished expression of CD19 in B-cell lymphomas.

BACKGROUND: CD19 is expressed on most B-cell lymphomas; however, the frequency and types of B-cell lymphomas with low-level expression of CD19 are not well characterized. METHODS: We reviewed flow cytometric histograms specifically for decreased CD19 expression on 349 cases analyzed by the Flow Cytometry Laboratory at University Hospitals of Cleveland (Cleveland, Ohio). Results of flow cytometry were correlated with the morphologic diagnosis. RESULTS: Of the cases reviewed, 125 (36%) showed a visible decrease in CD19 expression compared with normal B lymphocytes. Decreased CD19 expression was noted in 79% of follicular lymphomas (27 of 34), 36% of small lymphocytic lymphomas/chronic lymphocytic leukemias (82 of 228), 31% of mantle cell lymphomas (4 of 13), 24% of diffuse large B-cell lymphomas (8 of 33), and 13% of marginal zone B-cell lymphomas/lymphoplasmacytoid lymphomas (4 of 30) and was not observed in any Burkitt lymphoma (0 of 5) or hairy cell leukemia (0 of 6). Decreased CD19 expression was significantly more frequent in follicular lymphomas than in other lymphoma subtypes (P < 0.001). No significant difference was observed in the frequency of decreased CD19 expression based on histologic grade of follicular lymphoma. CONCLUSIONS: Diminished expression of CD19 expression occurs frequently in B-cell lymphomas, in particular follicular lymphoma, and may be helpful in identifying B-cell lymphoma cells in complex cell mixtures such as bone marrow specimens.

Adolescent↗
PubMed2005

Nonrandom aneuploidy of chromosomes 1, 5, 6, 7, 8, 9, 11, 12, and 21 induced by the benzene metabolites hydroquinone and benzenetriol.

The loss and gain of whole chromosomes (aneuploidy) is common in the development of leukemia and other cancers. In acute myeloid leukemia, the loss (monosomy) of chromosomes 5 and 7 and the gain (trisomy) of chromosome 8 are common clonal chromosomal abnormalities. Here, we have tested the hypothesis that metabolites of the human leukemogen benzene cause a higher rate of gain and loss among the chromosomes involved in leukemogenesis and, as such, are nonrandom and selective in their effects. Human peripheral blood was exposed to two metabolites of benzene, namely, hydroquinone (HQ) and benzenetriol (BT), and the ploidy status of nine different chromosomes (1, 5, 6, 7, 8, 9, 11, 12, and 21) was examined using fluorescence in situ hybridization of metaphase spreads. Poisson regression was used to provide interpretable incidence rate ratios and corresponding P values for all nine chromosomes. Statistically significant differences were found between the sensitivity of the nine chromosomes to gain or loss. Chromosomes 5 and 7 were highly sensitive to loss following HQ and BT exposure, whereas chromosomes 7, 8, and 21 were highly sensitive to gain in comparison to other chromosomes. Significant support for the a priori hypothesis that chromosomes 5 and 7 are more sensitive to loss induced by HQ and BT than the other seven chromosomes was also obtained. These data support the notion that benzene metabolites affect the ploidy status of specific chromosomes more than others and may initiate or promote leukemia induction through these specific effects.

Acute Disease↗
PubMed2005

Enhancement of cytomegalovirus detection in mink lung cells using CMV Turbo.

BACKGROUND: Detection of HCMV from clinical specimens was a slow process until the development of shell vial method with staining for immediate early antigen (IEA). Culture though still considered insensitive is widely used for other than blood samples. A method to improve culture sensitivity is desirable. OBJECTIVES: TurboTreat, a CMV pretreatment medium from Diagnostic Hybrids Inc., was evaluated on Mv1Lu, R-Mix and MRC-5 cells for improved sensitivity for HCMV detection. STUDY DESIGN: Monolayers of Mv1Lu, R-Mix and MRC-5 cells in 48-well plates were treated with TurboTreat for overnight (o/n), 4 h or left untreated and then inoculated with previously positive HCMV specimens. After o/n incubation, cells were fixed, stained and positive cells counted. RESULTS: CMV TurboTreat enhanced detection 2-3-fold after 4 h treatment and 4-6-fold after o/n treatment in Mv1Lu and R-Mix cells and to a lesser extend in MRC-5 cells with clinical isolates of HCMV. With frozen clinical specimens, Mv1Lu cells treated o/n, 4 h or untreated detected 23, 21 and 15 positive specimens, respectively. R-Mix cells detected 19, 18, and 14 positives, respectively and MRC-5 cells detected 16, 15 and 15 positives, respectively. In no case was a positive detected in another cell line regardless of treatment that was not detected in Mv1Lu treated o/n. CONCLUSION: The o/n pretreatment with CMV TurboTreat on Mv1Lu cells is the optimum condition of treatment and significantly enhanced the detection of HCMV. Even 4 h pretreatment of Mv1Lu cells showed significant enhancement over untreated Mv1Lu, R-Mix and MRC-5 cells. Pretreatment of Mv1Lu cells with CMV TurboTreat for 4 h or longer increased the sensitivity of rapid HCMV detection.

Animals↗
PubMed2005

Calcium and lanthanide affinity of the EF-loops from the C-terminal domain of calmodulin.

To obtain site-specific information about individual EF-hand motifs, the EF-hand Ca(2+)-binding loops from site III and site IV of calmodulin (CaM) were inserted separately into a non-Ca(2+)-binding cell adhesion protein, domain 1 of CD2 (denoted as CaM-CD2-III-5G-52 and CaM-CD2-IV-5G-52). Structural analyses using various spectroscopic methods have shown that the host protein CD2 retains its native structure after the insertion of the 12-residue loops. The Tb(3+) fluorescence enhancement upon formation of a Tb(3+)-protein complex and the direct competition by La(3+) and Ca(2+) suggest that native Ca(2+)-binding pockets are formed in both engineered proteins. Moreover, as revealed by NMR, both Ca(2+) and La(3+) specifically interact with the residues at the grafted EF-loop. The CaM-CD2-III-5G-52 has stronger affinities to Ca(2+), Tb(3+) and La(3+) than CaM-CD2-IV-5G-52, indicating differential intrinsic metal-binding affinities of the EF-loops.

Amino Acid Sequence↗
PubMed2005

Attenuation of ciclosporin-induced nephrotoxicity by dietary supplementation of seal oil in Sprague-Dawley rats.

Fish oil, rich in omega-3 (n-3) polyunsaturated fatty acids (PUFAs), has been reported to attenuate nephrotoxicity induced by ciclosporin (cyclosporine A). Harp seal oil is a rich source of n-3 PUFAs. This study investigated the ability of dietary seal oil to reduce nephrotoxicity caused by ciclosporin. Sprague-Dawley rats were maintained on a standard diet (with sunflower oil as lipid, SFO) or a diet enriched with seal oil (with 85% seal oil and 15% sunflower oil as lipid, SO) for four weeks before and four weeks after intravenous administration of ciclosporin (15 mg kg(-1) daily). Kidney function was assessed by measuring blood urea nitrogen, creatinine clearance, urinary N-acetyl-1-beta-D-glucosaminidase, 6-keto-prostaglandin F(1alpha), thromboxane B(2) and malondialdehyde. Systolic blood pressure (SBP) was monitored. Ciclosporin concentrations in blood were measured using liquid chromatographytandem mass spectrometry (LC-MS/MS). The fatty acid compositions of the diets and erythrocyte membranes were analysed by gas chromatography (GC). The results showed that nephrotoxicity was induced by ciclosporin in rats maintained on both SO and SFO diets. However, rats fed on SO diet endured less toxicity than those on SFO diet. The n-3 and n-6 PUFAs in the erythrocyte membrane of rats maintained on SO diet were found to be 10.79% and 11.93%, while those in rats maintained on SFO diet were found to be 1.67% and 22.71%, respectively. In conclusion, dietary supplementation of seal oil was found to reduce ciclosporin-induced nephrotoxicity in rats.

6-Ketoprostaglandin F1 alpha↗
PubMed2005

LC-MS determination and pharmacokinetic studies of ursolic acid in rat plasma after administration of the traditional chinese medicinal preparation Lu-Ying extract.

Sambucus chinensis L. is a native perennial herb distributed throughout China. In traditional Chinese medicine (TCM), this herb is known as Lu-Ying. Ursolic acid is the major effective constituent of Lu-Ying. A rapid, sensitive, and accurate liquid chromatography-mass spectrometry (LC-MS) method for the determination of ursolic acid in rat plasma was developed and validated. Plasma samples taken from rats that had received Lu-Ying extract orally were acidified with acetic acid and then extracted with a mixture of hexane-dichloromethane-2-propanol (20:10:1, v/v/v). Separation of ursolic acid was accomplished on a C(18) column interfaced with a single quadrupole mass spectrometer. The mobile phase consisting of methanol and water (95:5, v/v) was delivered at a flow rate of 1.0 ml/min. Atmospheric pressure chemical ionization was operated in negative-ion mode. Using selected ion-monitoring mode, the deprotonated molecules [M-H](-) at m/z 455 and 469 were used to quantify ursolic acid and glycyrrhetic acid (internal standard), respectively. The assay was shown to be linear over the range of 10-1000 ng/ml (r> or =0.9960) with a lower limit of quantification of 10 ng/ml. The method was shown to be reproducible and reliable with intraday precision below 7.8%, interday precision below 8.1%, accuracy within +/-4.3%, and mean extraction recovery excess of 83.6%, which were all calculated from the blank plasma sample spiked with ursolic acid at three concentrations of 20, 200, and 800 ng/ml. The LC-MS method has been successfully applied to pharmacokinetic studies of ursolic acid after oral administration of Lu-Ying ethanolic extract (at a dose containing 80.32 mg/kg ursolic acid) to rats. The main pharmacokinetic parameters were: t(1/2), 4.3 h; K(e), 0.16 1/h; t(max), 1.0 h; C(max), 294.8 ng/ml; AUC(0-t) and AUC(0-infinity), 1007.1 ng.h/ml and 1175.3 ng.h/ml, respectively.

Administration, Oral↗