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Biomedical subjects

Fei Wang

Publications and source records attributed to Fei Wang.

At least 127 records · Page 7Linked to original sources
PubMed2003

Trifluoperazine inhibits the MgATPase activity and in vitro motility of conventional and unconventional myosins.

Trifluoperazine, a calmodulin antagonist, has recently been shown to inhibit the MgATPase activity of scallop myosin in the absence of light chain dissociation (Patel et al. (2000) J Biol Chem 275: 4880-4888). To investigate the generality of this observation and the mechanism by which it occurs, we have examined the ability of trifluoperazine to inhibit the enzymatic properties of other conventional and unconventional myosins. We show that trifluoperazine can inhibit the actin-activated MgATPase activity of rabbit skeletal muscle myosin II heavy meromyosin (HMM), phosphorylated turkey gizzard smooth muscle myosin II HMM, phosphorylated human nonmuscle myosin IIA HMM and myosin V subfragment-1 (S1). In all cases half maximal inhibition occurred at 50-75 microM trifluoperazine while light chains (myosin II) or calmodulin (myosin V) remained associated with the heavy chains. In vitro motility of all myosins tested was completely inhibited by trifluoperazine. Chymotryptic digestion of baculovirus-expressed myosin V HMM possessing only two calmodulin binding sites yielded a minimal motor fragment with no bound calmodulin. The MgATPase of this fragment was inhibited by trifluoperazine over the same range of concentrations as the S1 fragment of myosin.

Actins↗
PubMed2003

Hepatocyte growth factor switches orientation of polarity and mode of movement during morphogenesis of multicellular epithelial structures.

Epithelial cells form monolayers of polarized cells with apical and basolateral surfaces. Madin-Darby canine kidney epithelial cells transiently lose their apico-basolateral polarity and become motile by treatment with hepatocyte growth factor (HGF), which causes the monolayer to remodel into tubules. HGF induces cells to produce basolateral extensions. Cells then migrate out of the monolayer to produce chains of cells, which go on to form tubules. Herein, we have analyzed the molecular mechanisms underlying the production of extensions and chains. We find that cells switch from an apico-basolateral polarization in the extension stage to a migratory cell polarization when in chains. Extension formation requires phosphatidyl-inositol 3-kinase activity, whereas Rho kinase controls their number and length. Microtubule dynamics and cell division are required for the formation of chains, but not for extension formation. Cells in the monolayer divide with their spindle axis parallel to the monolayer. HGF causes the spindle axis to undergo a variable "seesaw" motion, so that a daughter cells can apparently leave the monolayer to initiate a chain. Our results demonstrate the power of direct observation in investigating how individual cell behaviors, such as polarization, movement, and division are coordinated in the very complex process of producing multicellular structures.

Actins↗
PubMed2003

AAV-mediated expression of vascular endothelial growth factor induces choroidal neovascularization in rat.

PURPOSE: To develop a small-animal model of choroidal neovascularization (CNV) by injecting adeno-associated virus (AAV)-VEGF into the subretinal space (SRS) of rats. METHODS: An adeno-associated viral vector encoding human VEGF(165) was injected into the subretinal space (SRS) of Sprague-Dawley or Long Evans rats. Expression of VEGF was identified by RT-PCR and immunohistochemistry. Physiological and pathologic changes in the retina and choroid were evaluated by electroretinography, fluorescein angiography, light microscopy, and three-dimensional reconstruction of serial sections. RESULTS: Green fluorescent protein (GFP) and VEGF were expressed for at least 20 months in the retina and retinal pigment epithelium (RPE). Histologic sections showed extensive subretinal neovascularization, degenerating photoreceptors, and proliferating RPE at 5 weeks to 20 months after injection of AAV-VEGF. At 2 to 12 months after injection, leaking blood vessels were detected by fluorescein angiography. Electroretinogram a- and b-wave amplitudes were significantly decreased during this time. Three-dimensional reconstruction of serial sections demonstrated that choroidal blood vessels penetrated Bruch's membrane, one of them splitting into three branches in the SRS. In the current model, CNV was produced in 95% of the animals tested (19/20). It persisted for more than 20 months, a necessary requirement for modeling the development of CNV in age-related macular degeneration (AMD). CONCLUSIONS: In this study, a highly reproducible animal model of long-lasting CNV was developed. This model is being used to test antiangiogenic molecules to reduce or inhibit CNV and could be extended to primates.

Animals↗
PubMed2003

Production of soluble and functional engineered antibodies in Escherichia coli improved by FkpA.

Overproduction of genetically engineered antibodies, such as single-chain antibodies (scAbs) in Escherichia coli often results in insoluble and inactive products known as inclusion bodies. We now report that fusion or co-expression of FkpA, the E. coli periplasmic peptidyl-prolyl-isomerase with chaperone activity, substantially improves soluble and functional expression of scAbs. Anti-human bladder carcinoma scAb (PG) and anti-human CD3 x anti-human ovarian carcinoma-bispecific scAb (BH1) were fused with FkpA on the pTMF-based plasmid and expressed in E. coli. More than half of the amount of each expressed fusion protein FkpA-PG or FkpA-BH1 was soluble. In addition, the fusion protein cellulose-binding domain from Cellulomonas fimi (CBD)-PG and anti-human CD3 x anti-human CD28 x anti-human ovarian carcinoma-trispecific scAb (TRI) fused to the pelB (a signal peptide from pectate lysase B of a Bacillus sp.) signal sequence were co-expressed with FkpA under the control of the T7 promoter. A substantial portion of the co-expressed CBD-PG or TRI was soluble. Furthermore, PG, BH1, and TRI were biologically active as judged by ELISA and in vitro cytotoxicity assay. These results suggest that overexpression of FkpA should be useful in expressing heterologous proteins in E. coli.

Antibodies↗
PubMed2003

[Morphological observation of mesenchymal stem cells cultured with allogenic decalcified bone matrix].

OBJECTIVE: To investigate the feasibility of using decalcified bone matrix loaded with mesenchymal stem cells (MSCs) derived from adult human bone marrow as scaffolds for bone tissue engineering. METHODS: Allogenic decalcified bone matrix was prepared. MSCs were isolated from adult human bone marrow with monocyte-separating medium and then cultured in basic medium of mesenchymal stem cells (MSC). The third generation of the cultured cells were identified by FITC-antiCD105, seeded onto allogenic decalcified bone matrix followed by observation under scanning electron microscope and invert microscope a week later. RESULTS: Rapid proliferation of both primary and subcultured MSCs was observed, and CD105-positive cells reached 64.1% among the third generation of the cells. MSCs also grew well and proliferated rapidly after being cultured with decalcified bone matrix. CONCLUSION: Naive MSCs are ideal seeding cells for bone tissue engineering, and the allogenic decalcified bone matrix loaded with MSCs may be used as a good scaffold.

Bone Matrix↗
PubMed2003

[Growth factor and regeneration of intervertebral disc].

OBJECTIVE: To review research progress of the relation between growth factor and repair of intervertebral disc. METHODS: The recent articles on growth factor and repair of intervertebral disc were extensively reviewed. The expression of growth factor in intervertebral disc and the effect of growth factor on disc cells were investigated. RESULTS: Some growth factors play roles in the development and degeneration of intervertebral disc. Exogenous growth factor can increase proliferation of disc cells and production of proteoglycans and collagens. Gene of growth factor can be transferred to intervertebral disc cell by adenovirus. CONCLUSION: Growth factor plays an important role in the regulation of development and degeneration of interertebral disc. The above results show that the feasibility of usage of growth factor in the treatment of disc degeneration and in repair and reconstruction of disc.

Cell Division↗
PubMed2003

[Effect of IL-18 on peripheral blood monocytes from chronic hepatitis B patients].

OBJECTIVES: To explore the effect of IL-18 on peripheral blood monocytes (PBMCs) from chronic hepatitis B (CHB) patients and HBV DNA released by HepG2.2.15 cells, which were transfected with the gene of HBV. METHODS: PBMCs were isolated from 25 healthy persons and 25 CHB patients, which were co-cultured with HBcAg and IL-18 at different concentrations for 72 hours. The level of IFN-gamma in the culture supernatant of PBMCs was determined by ELISA. One patient' PBMCs were co-cultured for 96 hours with various concentrations of IL-18 and HepG2.2.15 cells which had been cultured for 24 hours, the supernatant was collected to detect HBV DNA level by PCR. RESULTS: When PBMCs were stimulated by HBcAg and IL-18 at various concentrations, the levels of supernatant IFN-gamma in the CHB group were much higher than those in the normal control group (at 0.2ng/ml: t=11.7, P<0.01; at 1.0ng/ml: t=16.19, P<0.01; at 5.0ng/ml: t=20.12, P<0.01), especially when the PBMCs were stimulated by HBcAg, IL-18 and IL-12 (1313.20pg/ml+-187.76pg/ml vs. 390.75pg/ml+-43.23pg/ml, t=23.94, P<0.01). The IFN-gamma level in the patients who were stimulated by HBcAg alone was much lower than the levels in the patients who were stimulated by HBcAg and IL-18 at various concentrations, and which were lower than those in the patients stimulated by HBcAg, IL-12 and IL-18 at the same concentrations (light: t=2.2, P<0.05; moderate: t=2.97, P<0.05). The HBV DNA content in the supernatant of co-cultivation with HepG2.2.15 cells and PBMCs was much higher than that of the two kinds of cells stimulated by HBcAg and IL-18 at various concentrations or HBcAg, IL-18 and IL-12/IFN-a1b. CONCLUSION: IL-18 can improve the PBMCs from CHB patients to produce a great deal of IFN-gamma, so it has a good application prospect in two aspects: immunoregulatory effects and increasing the ability to kill the cells infected with virus.

Adjuvants, Immunologic↗
PubMed2003

[Study on the determination of microelements in forest frog by inductively coupled plasma atomic emission spectroscopy].

In this paper, several trace elements in the forest frog samples were investigated by inductively coupled plasma atomic emission spectrometry (ICP-AES). During the experiment 5.00 mL HNO3 and 0.50 mL HClO4 were added to the sample, which then remained 24 hours before being heated to 120 degrees C and solved. This method can not only meet the requirement of ICP-AES, but also has many merits, such as simplicity, rapidity, precision and economy. The range of relative error was 0.17%-10.0% and the range of precision was 0.43%-9.96% for these elements.

Animals↗
PubMed2003

[Functional predation response of three main species to cotton bollworm in Xinjiang cotton-planting area].

Field insect population survey indicated that the principal natural predation enemies of cotton bollworm (Helicoverpa armigera Hubner) were Eringonidium graminicola Sundevall, Adonia variegata Goeze, Nabis sinoferus Hsiao. The results from experiments showed that the amount of prey consumed of these principal natural enemies increased when the density of prey rised, and decreased when the density of natural enemies increased. The predation function response fitted Holling's predation response equation II, and could be fit with the Disk Equation.

Animals↗
PubMed2003

[Karyotype analysis of Picris davurica chromosome].

OBJECTIVE: To systematically study Picris davurica Fisch chromosome karyotype. METHODS: Sections combined with micrograph were used to analyse chromosome. RESULTS: Chromosome number of normal diploid was 2n = 10, karyotype formula was K (2n) = 10 = 2 m + 8 sm, the relative length was 2n = 10 = 2 L + 2 M2 + 4 M1 + 2 S, which belonged to "3 A" type. The total length of chromosome groups was 12.51 microns, the total length of long arms was 8.36 microns, the AS. K% was 66.83%. The total volume of chromosome was 11.71 microns3. CONCLUSION: Picris davurica chromosome number and morphological character were clear.

Asteraceae↗
PubMed2003

[Clinical value of MR cholangiopancreatography combining with dynamic enhanced MRI in the detection of cholangiocarcinoma].

OBJECTIVES: To evaluate the clinical value of breath-hold magnetic resonance cholangiopancreatography (MRCP) combining with dynamic enhanced MRI in the diagnosis of cholangiocarcinoma. METHODS: MRCP findings of 88 cholangiocarcinoma patients proved surgically and pathologically were analyzed retrospectively. RESULTS: MRCP examination succeeded in all the 88 patients and the pancreaticobiliary ducts were shown satisfactorily. The accuracy of MRCP in the location of both hilar and extrahepatic cholangiocarcinoma was 100%, and the accuracy of detecting hilar and extrahepatic cholangiocarcinoma were 100% and 52.2%, respectively. Combining with dynamic enhanced MRI, the detecting accuracy of extrahepatic cholangiocarcinoma improved to 91.3%. CONCLUSIONS: MRCP examination has a high successful rate and can accurately determine the location of hilar and extrahepatic cholangiocarcinoma, and the accuracy of qualitative diagnosis for the former two is high. Combining with dynamic enhanced MRI, the specificity of determining extrahepatic cholangiocarcinoma is also high.

Adult↗
PubMed2002

The prepower stroke conformation of myosin V.

We have used electron microscopy and single-particle image processing to study head conformation in myosin V molecules. We find that in the presence of ATP, many heads have a sharply angled conformation that is rare in its absence. The sharply angled conformation is similar to a myosin II atomic structure proposed to mimic the prepower stroke state. The leading head in molecules attached to actin by both heads has a similar conformation, but is also sharply angled in a second plane by tethering through the trail head. The lead head lever joins the motor domain approximately 5 nm axially from where it joins the trail motor. These positions locate the converter subdomain and show the lead motor is in the prepower stroke conformation. Tethering by the trail head places the lead head motor domain at the correct axial position along the actin for binding, but at the wrong orientation. Attachment is achieved either by bending the lead head lever throughout its length or at the pliant point. The microscopy shows that most of the walking stride is produced by changes in lever angle brought about by converter movement, but is augmented by distortion produced by thermal energy.

Actins↗
PubMed2002

Phenotypic reversion or death of cancer cells by altering signaling pathways in three-dimensional contexts.

BACKGROUND: We previously used a three-dimensional (3D) reconstituted basement membrane (rBM) assay to demonstrate that tumorigenic HMT-3522 T4-2 human breast cells can be induced to form morphologically normal structures ("reversion") by treatment with inhibitors of beta1 integrin, the epidermal growth factor receptor (EGFR), or mitogen-activated protein kinase (MAPK). We have now used this assay to identify reversion and/or death requirements of several more aggressive human breast cancer cell lines. METHODS: Breast tumor cell lines MCF7, Hs578T, and MDA-MB-231 were cultured in 3D rBM and treated with inhibitors of beta1 integrin, MAPK, or phosphatidylinositol 3-kinase (PI3K). MDA-MB-231 cells, which lack E-cadherin, were transfected with an E-cadherin cDNA. The extent of reversion was assessed by changes in morphology and polarity, growth in 3D rBM or soft agar, level of invasiveness, and tumor formation in nude mice. RESULTS: All three cell lines showed partial reversion (MCF7 the greatest and Hs578T the least) of tumorigenic properties treated with a single beta1 integrin, MAPK, or PI3K inhibitor. Combined inhibition of beta1 integrin and either PI3K or MAPK resulted in nearly complete phenotypic reversion (MDA-MB-231, MCF7) or in cell death (Hs578T). E-cadherin-transfected MDA-MB-231 cells showed partial reversion, but exposure of the transfectants to an inhibitor of beta1 integrin, PI3K, or MAPK led to nearly complete reversion. CONCLUSION: The 3D rBM assay can be used to identify signaling pathways that, when manipulated in concert, can lead to the restoration of morphologically normal breast structures or to death of the tumor cells, even highly metastatic cells. This approach may be useful to design therapeutic intervention strategies for aggressive breast cancers.

Animals↗
PubMed2002

Mutations in human nonmuscle myosin IIA found in patients with May-Hegglin anomaly and Fechtner syndrome result in impaired enzymatic function.

A family of autosomal-dominant diseases including May-Hegglin anomaly, Fechtner syndrome, Sebastian syndrome, Alport syndrome, and Epstein syndrome are commonly characterized by giant platelets and thrombocytopenia. In addition, there may be leukocyte inclusions, deafness, cataracts, and nephritis, depending on the syndrome. Mutations in the human nonmuscle myosin IIA heavy chain gene (MYH9) have been linked to these diseases. Two of the recently described mutations, N93K and R702C, are conserved in smooth and nonmuscle myosins from vertebrates and lie in the head domain of myosin. Interestingly, the two mutations lie within close proximity in the three-dimensional structure of myosin. These two mutations were engineered into a heavy meromyosin-like recombinant fragment of nonmuscle myosin IIA, which was expressed in baculovirus along with the appropriate light chains. The R702C mutant displays 25% of the maximal MgATPase activity of wild type heavy meromyosin and moves actin filaments at half the wild type rate. The effects of the N93K mutation are more dramatic. This heavy meromyosin has only 4% of the maximal MgATPase activity of wild type and does not translocate actin filaments in an in vitro motility assay. Biochemical characterization of the mutant is consistent with this mutant being unable to fully adopt the "on" conformation.

Base Sequence↗
PubMed2002

Comparison of two in vitro methods for the measurement of recombinant human TSH bioactivity.

Thyroid stimulating hormone (TSH), a pituitary glycoprotein hormone, is a potent inducer of intracellular cAMP production. Two methods for measuring TSH bioactivity were evaluated and compared. One assay is based on using a radioimmunoassay (RIA) to measure the recombinant human TSH-induced increase in cAMP using a bovine thyroid membrane isolate. The other is based on a Chinese hamster ovary (CHO) cell line that has been transfected with the TSH receptor and a cAMP-responsive luciferase reporter. The within-assay coefficient of variation for the membrane-based assay was determined to be approximately 35% compared with approximately 25% for the cell-based assay. Twenty-one preparations of recombinant human TSH (rhTSH) were tested using both methods. No significant difference was detected between the data sets and no assay bias was present. Both assay systems provide a suitable means for measuring the activity of rhTSH. The advantage of the membrane-based assay is the relatively small quantity of TSH needed for analysis. However, the average time required to analyse a sample using the membrane-based method was more than twice as long as that needed to test a sample in the cell-based assay. Other advantages of the cell-based method include the use of a 96-well format, which facilitates the analysis of several concentrations of rhTSH within one assay plate, and the use of a non-radioactive endpoint.

Animals↗