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Biomedical subjects

S Feng

Publications and source records attributed to S Feng.

At least 109 records · Page 6Linked to original sources

Identification of carbohydrates on the surface membrane of pathogenic and nonpathogenic piscine haemoflagellates, Cryptobia salmositica, C. bullocki and C. catostomi (Kinetoplastida).

Carbohydrates and protein glycoconjugates on the cell membranes of Cryptobia salmositica, C. bullocki and C. catostomi were analyzed using 13 highly purified lectins (unlabelled or digoxigenin/biotin labelled). No agglutinations were observed with C. salmositica, C. bullocki and C. catostomi using lectin TPA (Tetragonolobus purpureas agglutinin, for alpha-L-fucose). C. salmositica was agglutinated by 3 of 12 lectins [Con A, for alpha-man and alpha-D-glc; PSA, for alpha-man; PWM, for (glcNAc)3], while C. bullocki was agglutinated by 8 lectins and C. catostomi was agglutinated by 10 lectins. Glycoconjugate analysis with digoxigenin or biotin labelled lectins showed a species-specific staining pattern in pathogenic and nonpathogenic Cryptobia spp. The nonpathogenic C. catostomi had the strongest reaction. These results indicate that the surface carbohydrate residues and glycoconjugate compositions on Cryptobia spp. are different between species they may be related to the virulence of the parasite.

Agglutination↗

Inhibition of growth of malignant rat prostate tumor cells by restoration of fibroblast growth factor receptor 2.

A loss of expression of fibroblast growth factor (FGF) receptor 2 IIIb (FGFR2IIIb), which responds to stroma-derived FGF, accompanies progression of premalignant androgen-responsive rat prostate tumor epithelial cells to the malignant phenotype. Concurrently, the level of FGFR2 gene expression is reduced and lost altogether in over 30% of cells, whereas all malignant cells abnormally express FGFR1, which is normally confined to stromal cells (S. Feng et al., Cancer Res., 57:5369-5378, 1997). To determine the relative roles of the FGFR2 and FGFR1 kinases in growth of malignant cells, we transfected malignant prostate epithelial cells with the wild-type FGFR2IIIb kinase and an artificial chimeric construct (FGFR2IIIb/R1) composed of the FGFR2IIIb ectodomain and the FGFR1 kinase domain. Population growth kinetics, in both the absence and presence of FGF-7, which binds only the FGFR2IIIb ectodomain, were then examined in the transfected cell populations. In contrast to the untransfected malignant tumor cells and those expressing the FGFR2IIIb/R1 chimera, FGF-7 caused a dose-dependent net inhibition of the population growth rates of cells expressing the full-length FGFR2IIIb kinase. The results suggest that although the FGFR2 kinase can mediate positive mitogenic effects, it mediates a net restriction on the growth of prostate tumor epithelial cells relative to FGFR1. Highly malignant prostate tumor cells, which have lost the FGFR2 tyrosine kinase, retain the cellular response mechanisms to it. Restoration of the FGFR2 kinase to malignant tumors that are refractory to treatment may present a new avenue for gene therapy.

Animals↗

Characterization of a 200 kDa glycoprotein (Cs-gp200) on the pathogenic piscine haemoflagellate Cryptobia salmositica.

The 200 kDa antigenic doublet of the pathogenic haemoflagellate Cryptobia salmositica Katz, 1951, was detected using a monoclonal antibody (MAb-001) in 1-dimensional SDS-PAGE. This antigenic doublet is a glycoprotein since it was susceptible to both protease K and to sodium m-periodate oxidation and is designated Cs-gp200. It has a PI (isoelectric point) value of about 5.5 (using 2-dimensional gel electrophoresis). It migrated faster under reducing conditions than under nonreducing conditions and was partitioned into the aqueous phase in Triton X-114 phase separation. It is a secretory-excretory product since it was detected in a non-protein culture medium with C. salmositica. These results suggest that the Cs-gp200 is a glycoprotein consisting of carbohydrate determinants and conformational polypeptide with internal disulphide bonds. It is a hydrophilic antigen, is a secretory product of the parasite, and plays a important role in antibody elicitation in immunized fish.

Animals↗

Direct activation of inward rectifier potassium channels by PIP2 and its stabilization by Gbetagamma.

Inward rectifier K+ channels, which modulate electrical activity in many cell types, are regulated by protein kinases, guanine-nucleotide-binding proteins (G proteins) and probably actin cytoskeleton. Generation of phosphatidylinositol 4,5-bisphosphate (PIP2) by ATP-dependent lipid kinases is known to activate inward rectifier K+ channels in cardiac membrane patches. Here we report that several cloned inward rectifier K+ channels directly bind PIP2, and that this binding correlates with channel activity. Application of ATP or PIP2 liposomes activates the cloned channels. Stabilized by lipid phosphatase inhibitors, PIP2 antibodies potently inhibit each channel with a unique rate (GIRK1/4 approximately GIRK2 >> IRK1 approximately ROMK. Consistent with the faster dissociation of PIP2 from the GIRK channels, the carboxy terminus of GIRK1 binds 3H-PIP2 liposomes more weakly than does that of IRK1 or ROMK1. Mutation of a conserved arginine to glutamine at position 188 reduces the ability of ROMK1 to bind PIP2 and increases its sensitivity to inhibition by PIP2 antibodies. Interactions between GIRK channels and PIP2 are modulated by the betagamma subunits of the G protein (Gbetagamma). When GIRK1/4 channels are allowed to run down completely, they are not activated by addition of Gbetagamma alone, but application of PIP2 activates them in minutes without Gbetagamma and in just seconds with Gbetagamma. Finally, coexpression of Gbetagamma with GIRK channels slows the inhibition of K+ currents by PIP2 antibodies by more than 10-fold. Thus Gbetagamma activates GIRK channels by stabilizing interactions between PIP2 and the K+ channel.

Amino Acid Sequence↗

Electrophoretic affinity chromatography: method validation.

A new method for preparative-scale separation of biomolecules, electrophoretic affinity chromatography (EAC), is proposed in this paper. Separation by EAC is carried out in a long and ribbon-like multicompartment electrolyser separated by membranes, in which the two central compartments are used for packing the gel matrix and for sample loading respectively. Next to the central compartments are the elution compartments and electrode compartments. The electric field is applied perpendicular to the fluid flow in the compartments. Adsorption and desorption steps may both be carried out in the presence of an electric field, which transports the target components into the gel compartment for adsorption and the impurities into the elution compartments for washing. After the adsorption step an elution solution is introduced and the product is released from the gel matrix and washed out. Separation of human serum albumin (HSA) from human serum gives HSA product of high purity, as demonstrated by isoelectric focusing analysis. The characteristics of electrophoretic binding of HSA on Blue Sepharose Fast Flow are examined. The preliminary results show that this new method has advantages in terms of high rate of mass transfer and ease of scaling up, which are of particular interest when large-scale separation of biomolecules is considered.

Adsorption↗

Biochemical characterisation of an epitope on the surface membrane antigen (Cs-gp200) of the pathogenic piscine haemoflagellate Cryptobia salmositica Katz 1951.

A protective surface antigen (200 kDa) on C. salmositica was detected using a monoclonal antibody (mAb-001). Enzymatic studies on the epitope indicated that it was sensitive to nonspecific protease K and to site-specific trypsin and protease V8 but not to alpha-chymotrypsin. The reactivity of the epitope with mAb-001 was not affected when the antigen was denatured with 8 M urea; however, reduction of the antigen with dithiothreitol destroyed the epitope. The epitope was susceptible to sodium m-periodate oxidation and N-glycosidase F, but not to O-glycosidase or neuraminidase. It was also sensitive to mild potassium hydrochloride hydrolysis and to phospholipase C, which is specific for phosphatidylinositol. These results suggest that the epitope consists of a polypeptide, a carbohydrate, and probably a phospholipid. The asparagine-bound N-glycosidically linked hybrid-type carbohydrate chain has the minimum length of a chitobiose core unit. There is probably a phosphatidylinositol residue which anchors the polypeptide to the surface membrane. The antigen is extensively posttranslationally modified.

Amidohydrolases↗

Granulocytic ehrlichiosis in the laboratory mouse.

C3H mice that were inoculated with ehrlichiae isolated from a patient with human granulocytic ehrlichiosis (HGE) developed anemia and leukopenia, but by day 24, they returned to normal values. Granulocytic morulae were present in peripheral blood and spleen smears on days 5 and 10, and there was a reduction in morulae on day 17. Ehrlichiae were present in HL-60 cell cultures of blood and spleen from all mice at all intervals. Pathogenicity, but not infectivity, waned with mouse passage but could be resurrected by SCID mouse passage. Various methods were tested for their relative sensitivity in detecting infection: blood smears, HL-60 cell cultures, polymerase chain reaction (PCR) amplification of a 16S recombinant DNA target, and a mouse infectivity assay. All assays detected the HGE agent in blood during early infection, but PCR and the mouse infectivity assay were most sensitive during late infection. Xenodiagnosis demonstrated that mice remain persistently infected through 55 days.

Age Factors↗

Humoral immunity to Borrelia burgdorferi N40 decorin binding proteins during infection of laboratory mice.

A Borrelia burgdorferi N40 genomic expression library was screened with serum from actively infected mice to identify gene products that elicit protective immunity. A clone that contained a putative bicistronic operon containing two genes that encoded 20- and 22-kDa lipoproteins was identified and sequenced. These genes showed homology with the genes encoding decorin binding proteins DbpB and DbpA, respectively, of B. burgdorferi 297 and B31. N40-dbpA DNA hybridized with B. burgdorferi N40 DNA on a single 48-kb linear plasmid. Homologous genes could be amplified under various degrees of stringency by PCR or hybridized by Southern blotting from B. burgdorferi sensu stricto N40 and B31, and from B. burgdorferi sensu lato PBi and 25015, but not PKo. Recombinant N40-DbpB and N40-DbpA were reactive with antibody in serum from infected mice, and serum was more reactive against N40-DbpA than against B. burgdorferi N40 recombinant P39, OspC, or OspA. Sera from mice infected with B. burgdorferi sensu lato strains PKo and PBi were weakly reactive against N40-DbpB and N40-DbpA, and sera from mice infected with 25015 were moderately reactive, compared to sera from mice infected with B. burgdorferi N40. Hyperimmunization of mice with N40-DbpA, but not N40-DbpB, induced protective immunity against syringe challenge with cultured B. burgdorferi N40. DbpA may therefore be one of the antigens responsible for eliciting protective antibody known to exist in serum from infected mice. DNA amplification and serology suggest that DbpB and DbpA are likely to have homologs throughout the B. burgdorferi sensu lato family, but they are likely to be heterogeneous.

Adhesins, Bacterial↗

Acquisition and transmission of the agent of human granulocytic ehrlichiosis by Ixodes scapularis ticks.

The purpose of the present study was to investigate the transmission of a human isolate of the agent of human granulocytic ehrlichiosis (HGE agent) from infected mice to larval ticks and to examine the population kinetics of the HGE agent in different stages of the tick life cycle. The HGE agent was quantitated by competitive PCR with blood from infected mice and with Ixodes scapularis ticks. The median infectious dose for C3H mice was 10(4) to 10(5) organisms when blood from an infected severe combined immunodeficient mouse was used as an inoculum. Uninfected larval ticks began to acquire infection from infected mice within 24 h of attachment, and the number of HGE agent organisms increased in larval ticks during feeding and after detachment of replete ticks. Molted nymphal ticks, infected as larvae, transmitted infection to mice between 40 and 48 h of attachment. Onset of feeding stimulated replication of the HGE agent within nymphal ticks. These studies suggest that replication of the HGE agent during and after feeding in larvae and during feeding in nymphs is a means by which the HGE agent overcomes inefficiencies in acquisition of infection by ticks and in tick-borne transmission to mammalian hosts.

Animals↗

In vivo and in vitro modulation of immune stimulatory capacity of primary dendritic cells by adenovirus-mediated gene transduction.

Dendritic cells (DCs) are potent antigen-presenting cells which are key leukocytes in the initiation of cell-mediated organ graft rejection, antiviral immunity, and antitumor responses. In this study we demonstrate that genetic modification of primary human and mouse DCs by adenoviral gene transfer is an effective means of induction and modulation of antigen presentation by DCs. An adenovirus vector (AdLacZ) was used to express an intracellular model antigen beta-galactosidase (beta-gal) in DCs. Our results show that 30-40% of precursor dendritic cells (PDCs) derived from human umbilical cord blood and circulating mature blood DCs express high levels beta-galactosidase (beta-gal) after infection with AdLacZ with no cytopathic effect observed. In vitro, AdLacZ transduced PDCs and DCs demonstrated a 10- to 20-fold higher mixed lymphocyte reaction (MLR) stimulatory capacity as compared to that of monocytes. In vivo, immunization with AdLacZ transduced mouse DCs resulted in more potent cytotoxic T lymphocyte (CTL) responses against the predicted H-2 restricted beta-gal epitope as compared to CTL responses obtained by beta-gal peptide pulsed DCs. Modulations of the MLR stimulatory capacity of DCs were examined by expression of mouse B7 and CTLA-4Ig. The results show that expression of mouse B7 by a recombinant adenoviral vector (Ad7) significantly enhances the MLR stimulatory capacity of human DCs. In contrast, expression of CTLA-4Ig (AdCTLA-4Ig) reduces the MLR stimulatory capacity of the transduced cells. We conclude that recombinant adenovirus can readily be used for genetic modulation DC-induced immune responses in vivo and in vitro. DCs targeted for induction of specific antigen responses or for modulation of the immune stimulatory capacity may have a potential use in the control of transplantation rejection or viral infections.

Adenoviridae↗

[Effects of mifepristone on the serum nitric oxide level and uterine blood flow in early pregnant women].

OBJECTIVE: To study the effects of mifepristone on serum nitric oxide (NO) level and uterine blood flow in early pregnant women. METHODS: Serum NO concentrations of 58 early pregnant women and 15 normal nonpregnant women were measured by a copper-coated cadmium reduction method, and uterine artery and trophoblastic blood flow were determined by color doppler flow imaging (CDFI). The ultrastructure of villi and decidua were studied under electron microscopy. RESULTS: Compared with nonpregnant women, serum NO concentration was significantly higher in early pregnant women(P < 0.01). After administration of mifepristone, serum NO level was significantly decreased, resistance index(RI) and systolic to diastolic ratio(S/D) ratio of uterine artery and trophoblastic blood flow were significantly higher than that of normal early pregnancy(P < 0.05). CONCLUSION: Mifepristone may exert effects on uterine blood flow by decreasing serum NO level.

Abortifacient Agents, Steroidal↗

[Outcome of autologous bone marrow transplantation in 73 patients with acute leukemia].

OBJECTIVE: To evaluate the clinical outcome of autologous bone marrow transplantation(ABMT) in acute leukemias. METHODS: Data of 73 acute leukemia patients performed ABMT in our hospital from October 1986 to December 1997 were retrospectively analyzed. The median age was 25.5(9-48) years. Forty-three of them were ANLL [CR1 35, CR2 or early relapse (ER)8], 30 were ALL (CR1 25, CR2 or ER 5). Conditioning regimens were CTX 120 mg/kg + STBI 9-10 Gy or Bu 16 mg/kg or Mel 160-180 mg/m2 + Ara-C 4 g/m2. RESULTS: All patients engrafted successfully. The median follow-up duration was 806 (32-3400) days. The 3-year probabilities of disease-free survival (DFS) for ANLL and ALL in CR1 were 54.9 +/- 7.7% and 67.0% +/- 10.6%, respectively, and the probabilities of relapse were 39.3% +/- 9.3% and 23.7% +/- 10.6%, respectively. CONCLUSION: To decrease relapse and increase DFS, patients with acute leukemia in CR1 who have no HLA-matched related donor are recommended for ABMT.

Adolescent↗

[Synthesis and anti-Candida albicans properties of L-4-oxalysine-N3-4-methoxyfumaroyl-L-2,3-diaminopropanoic-containing peptide analogues].

In order to improve the inhibitory activity of L-4-oxalysine(abbreviated as I-677) against clinically important pathogen Candida albicans, double-warheads peptide analogues containing I-677 and N3-4-methoxyfumaroyl-L-2,3-diaminopropanoic acid(FMDP) were designed based on the concept of "Illicit Transport" and peptide transport specificities of C. albicans. One compound of I-677-FMDP and seven compounds of I-677-AA-FMDP(AA = Nva, Leu, Val, Phe, Pro, D-Pgly, D, L-p-Cl-Phe) were synthesized and examined for antifungal activities. The results of anti-Candida albicans test in vitro of these double- warheads peptide analogues containing I-677 and FMDP showed very high activities against Candida albicans. The molar MIC (molar minimum inhibitory concentration) ratio of free I-677 to I-677-AA-FMDP is 40-770 and that of FMDP is 60-1130. The values of molar MIC of I-677-AA-FMDP varied from 6.56 x 10(-9) mol.disk-1 to 3.5 x 10(-10) mol.disk-1. The results of competitive antagonism studies indicated that I-677-AA-FMDP were transported into Candida albicans cells by the di-tripeptide permease and subsequently hydrolyzed by intracellular peptidases, releasing free I-677 and FMDP inside cells. The result of antienzymic degradation test in vitro of I-677-FMDP showed that compound I-677-FMDP was resistant to the hydrolysis by carboxypeptidase A.

Animals↗

GC/MS analysis of m-hydroxybenzoylecgonine in urine. Forensic implication in cocaine use.

m-Hydroxybenzoylecgonine (m-OH-BE) and d3-m- hydroxybenzoylecgonine (d3-m-OH-BE) have been synthesized, and a GC/MS procedure with d3-m-OH-BE as internal standard has been developed. Among 24 human urine specimens that were positive for BE, all of them have shown detectable levels of m-OH-BE with 75% of the specimens exceeding the LoQ (5 ng/mL), compared with only 50% of the specimens containing detectable levels of EME. The presence of m-OH-BE in urine suggested that this metabolite may serve as a valuable marker of cocaine use in addition to BE and EME.

Cocaine↗

Fibroblast growth factor receptor 2 limits and receptor 1 accelerates tumorigenicity of prostate epithelial cells.

Progressive loss of the differentiated phenotype and communication with stroma accompanies the transition of nonmalignant rat prostate epithelial cells to anaplastic, malignant tumors. Here we show that cell surface expression of the fibroblast growth factor receptor 2 (FGFR2) tyrosine kinase is reduced in malignant tumor cell populations (type II) and undetectable at the mRNA level in 30% of cells. This is in addition to the irreversible loss by splice switching of the FGFR2 ectodomain that abrogates response to FGF-7 and homologues from the stroma. One hundred % of type II malignant cells express FGFR1, which is normally expressed in the stroma. Expression of the FGFR1 kinase in premalignant type I tumor epithelial cells by transfection accelerated progression to the malignant phenotype. In contrast to the FGFR2 kinase fused to the ectodomain of FGFR1, the FGFR1 kinase failed initially to support a mitogenic response to FGF-2 in type I tumor cells. However, the FGFR1-transfected cells acquired a mitogenic response after extensive proliferation of the cell population. Resident FGFR2 and ectopic FGFR1 appeared to be partitioned in the type I cells, because neither full-length nor truncated isoforms of FGFR1 affected the mitogenic response of the other. Restoration of the FGFR2IIIb kinase to malignant cells expressing FGFR1 depressed tumor growth rates, restored responsiveness to stromal cells, and restored epithelial cell differentiation. These observations reveal that homologous FGFR1 and FGFR2 kinases play very different roles in cell growth and differentiation and in development and support of the malignant phenotype.

Animals↗

A homeo-interaction sequence in the ectodomain of the fibroblast growth factor receptor.

Interaction of fibroblast growth factor receptors (FGFR) sufficient for a trans-phosphorylation event in which one intracellular domain is substrate for the other is essential for signal transduction. By analysis of the direct interaction of recombinant constructions co-expressed in baculoviral-infected insect cells, we identified a 17-amino acid sequence that is required for the stable interaction between ectodomains of FGFR. The sequence 160ERSPHRPILQAGLPANK176 (Glu160-Lys176) connects immunoglobulin modules II and III. In insect cells, the interaction between Glu160-Lys176 domains occurs independently of intact heparin or FGF binding domains. The sequence is not required for the binding of heparin or FGF-1, but is essential for mitogenic activity of the FGFR kinase in mammalian cells. The results support a model in which the homeo-interaction between Glu160-Lys176 in the ectodomain contributes to the interaction between intracellular domains in mammalian cell membranes (Kan, M., Wang, F., Kan, M., To, B., Gabriel, J. L., and McKeehan, W. L. (1996) J. Biol. Chem. 271, 26143-26148). We propose that the Glu160-Lys176 domain plays a pivotal role in restriction of the interaction between kinases by pericellular matrix heparan sulfate proteoglycan and divalent cations. Restrictions are overcome by FGF or constitutively by diverse gain of function mutations which cause skeletal and craniofacial abnormalities.

Amino Acid Sequence↗

Regulatory intramolecular association in a tyrosine kinase of the Tec family.

The T-cell-specific tyrosine kinase Itk is a member of the Tec family of non-receptor tyrosine kinases, and is required for signalling through the T-cell antigen receptor (TCR). The role of Itk in TCR signalling and the manner in which Itk activity is regulated are not well understood. Substrate binding and enzymatic activity of the structurally related Src kinases are regulated by an intramolecular interaction between the Src-homology-2 (SH2) domain and a phosphotyrosine. Although Itk also contains SH3, SH2 and tyrosine kinase domains, it lacks the corresponding regulatory phosphorylation site, and therefore must be regulated by an alternative mechanism. The proline-rich sequence adjacent to the SH3 domain of Tec family kinases contains an SH3 ligand, potentially allowing a different intramolecular interaction. By using multidimensional nuclear magnetic resonance we have determined the structure of a fragment of Itk, confirming that these domains interact intramolecularly. Formation of this intramolecular SH3-ligand complex prevents the Itk SH3 domain and proline-rich region from interacting with their respective protein ligands, Sam68 and Grb-2. We believe that this structure represents the first example of an intramolecular interaction between an SH3 domain and a proline-rich ligand, and has implications for the regulation of Tec family kinases.

Amino Acid Sequence↗