Perspectives. Insurance reform sleeper: Medicare contracting overhaul.
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Centers for Medicare and Medicaid Services, U.S.↗
Biomedical subjects
S Williams
Publications and source records attributed to S Williams.
Bistratene A causes phosphorylation of talin and redistribution of actin microfilaments in fibroblasts: possible role for PKC-delta.
Bistratene A is a marine toxin which induces phosphorylation of cellular proteins. Our current evidence indicates that this occurs through activation of protein kinase C-delta. In fibroblasts bistratene A causes rounding up of the cells and a rapid disappearance of vinculin staining and actin stress fibers as detected by fluorescence immunohistochemistry. Phosphorylation of the focal adhesion protein, talin, is increased after bistratene A treatment and this is inhibited by calphostin C, a specific inhibitor of PKC. No changes in the phosphorylation status of vinculin, tubulin, or vimentin were observed in the presence of the toxin. Treatment with bistratene A caused a redistribution of PKC-delta from cytosolic and membrane compartments to the nuclear fraction. There was no effect on the subcellular distribution of any other PKC isoform. These results demonstrate that phosphorylation of talin is implicated in the disruption of actin microfilaments in fibroblasts by bistratene A and that this is most likely mediated by PKC-delta.
Acetamides↗
Regulation of the Lck SH2 domain by tyrosine phosphorylation.
Src homology 2 (SH2) domains bind to phosphotyrosine (Tyr(P)) residues in specific sequence contexts in other proteins and thereby mediate tyrosine phosphorylationdependent protein-protein interactions. The SH2 domain of the Src family kinase Lck is phosphorylated at tyrosine 192 in T cells upon T cell antigen receptor triggering. We have studied the consequences of this phosphorylation on the properties of the SH2 domain and on the function of Lck in T cell activation. We report that phosphorylation at Tyr192 reduced the capacity of the isolated SH2 domain to bind a high affinity peptide ligand and Tyr(P)-containing cellular proteins. This effect was mimicked by mutation of Tyr192 to an acidic residue. In intact T cells, where Lck participates in T cell antigen receptor signal transduction in an SH2 domain-dependent manner, phosphorylation of Tyr192 correlated with reduced downstream signaling. Our results indicate that tyrosine phosphorylation of the SH2 domain of Lck terminates its high affinity binding to ligands, thereby negatively regulating its participation in T cell antigen receptor signaling. This represents a novel mechanism for the regulation of the function of SH2 domains.
Amino Acid Sequence↗
Identification of the site in the Syk protein tyrosine kinase that binds the SH2 domain of Lck.
The Syk protein tyrosine kinase (PTK) is expressed in many hematopoietic cells and is involved in signaling from various receptors for antigen and Fc portions of IgG and IgE. Upon cross-linking of these receptors, Syk is rapidly phosphorylated on tyrosine residues and enzymatically activated. We and others have found that the Lck kinase, a member of the Src family of PTKs, binds through its Src homology (SH) 2 domain to tyrosine phosphorylated Syk and to the related Zap kinase. Here we report that this interaction is direct and identify the two tandem tyrosines at the autophosphorylation site of Syk, Tyr518, and Tyr519, as the binding site for the SH2 domain of Lck. Mutation of either or both tyrosines to phenylalanines abrogated binding, while mutation of a second repetition of the motif at Tyr539 and Tyr540, or of the three tyrosines in the C terminus of Syk, did not. The SH2 domain of Lck bound the autophosphorylation site only when both Tyr518 and Tyr519 were phosphorylated. In intact cells the binding of the SH2 domain of Lck correlated with the ability of Syk to induce tyrosine phosphorylation of cellular proteins.
Amino Acid Sequence↗
Perspectives. Cornerstone to reform: consumer satisfaction surveys.
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Consumer Advocacy↗
Regional localization of over 300 loci on human chromosome 22 using a somatic cell hybrid mapping panel.
A somatic cell hybrid panel, consisting of 25 cell lines, has been developed to localize loci subregionally on chromosome 22. Over 300 markers in the form of STSs or hybridization probes have been assigned to one of 24 unique regions or "bins" using this panel. This ordered collection of markers will aid in the assembly of physical maps and contigs of chromosome 22 and assist in positional cloning of disease loci mapped to chromosome 22.
Animals↗
Involvement of Src-homology-2-domain-containing protein-tyrosine phosphatase 2 in T cell activation.
Activation of resting T lymphocytes by ligands to the complex of T cell antigen receptor (TCR) and CD3 is initiated by a series of critical tyrosine phosphorylation and dephosphorylation events. Protein-tyrosine kinases of the Syk, Src and Csk families and the CD45 protein-tyrosine phosphatase (PTPase) are known to be involved in these early biochemical reactions. We have found that one of the two T-cell-expressed SH2-domain-containing PTPases, SHPTP2, is rapidly phosphorylated on tyrosine upon addition of anti-CD3 mAbs. This response was absent in cells lacking the Src family kinase Lck. Concomitantly with tyrosine phosphorylation, SHPTP2 co-immunoprecipitated with two unphosphorylated cellular proteins; phosphatidylinositol 3-kinase p85 and Grb2. Binding of SHPTP2 to Grb2 occurred through the SH2 domain of Grb2, while the association between SHPTP2 and p85 seemed to be mediated through Grb2 as an intermediate. In addition, many other molecules associate with Grb2 and may thereby become juxtaposed to SHPTP2. Our results indicate that SHPTP2 participates actively at an early stage in TCR signaling and that its phosphorylation on tyrosine may direct a Grb2-dependent association with selected substrates.
Adaptor Proteins, Signal Transducing↗
Pearson's correlation coefficient.
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Analysis of Variance↗
Human immunodeficiency virus infection of bone marrow endothelium reduces induction of stromal hematopoietic growth factors.
The majority of human immunodeficiency virus (HIV)-seropositive patients develop bone marrow abnormalities associated with hematopoietic malfunction during the progression of disease. One important manifestation of HIV-associated hematopoietic dysfunction is that after myelosuppression, bone marrow recovery, a process known to be mediated in part by the production of stromal cell-derived hematopoietic growth factors, is impaired. We sought to test the hypothesis that bone marrow stromal cells are infected by HIV-1 in vivo and that production of certain stromal cell-derived hematopoietic growth factors is deficient as a consequence. In this report, we demonstrate that bone marrow microvascular endothelial cells (MVEC), a key element of the stroma, are the predominant cells infected by HIV (5% to 20%) in bone marrow stromal cultures obtained from 11 consecutive HIV-seropositive patients. Although HIV-infected stromal cultures enriched for MVEC constitutively express normal levels of interleukin (IL)-4, IL-6, granulocyte (G)-colony-stimulating factor (CSF), granulocyte-macrophage (GM)-CSF, tumor necrosis factor (TNF)-alpha, transforming growth factor (TGF)-beta, and Steel factor, IL-1 alpha-induced release of IL-6 and G-CSF is significantly reduced in these cultures. These observations suggest that HIV infection of bone marrow MVEC reduces the capacity of hematopoietic stroma to respond to regulatory signals that normally augment blood cell production during periods of increased demand.
Adult↗
Fast events in protein folding: helix melting and formation in a small peptide.
The helix is a common secondary structural motif found in proteins, and the mechanism of helix-coil interconversion is key to understanding the protein-folding problem. We report the observation of the fast kinetics (nanosecond to millisecond) of helix melting in a small 21-residue alanine-based peptide. The unfolding reaction is initiated using a laser-induced temperature jump and probed using time-resolved infrared spectroscopy. The model peptide exhibits fast unfolding kinetics with a time constant of 160 +/- 60 ns at 28 degrees C in response to a laser-induced temperature jump of 18 degrees C which is completed within 20 ns. Using the unfolding time and the measured helix-coil equilibrium constant of the model peptide, a folding rate constant of approximately 6 x 10(7) s-1 (t1/2 = 16 ns) can be inferred for the helix formation reaction at 28 degrees C. These results demonstrate that secondary structure formation is fast enough to be a key event at early times in the protein-folding process and that helices are capable of forming before long range tertiary contacts are made.
Circular Dichroism↗
Identification of a human erythroid progenitor cell population which expresses the CD34 antigen and binds the plant lectin Ulex europaeus I.
Two and three color flow cytometry of normal human bone marrow was used to identify CD34+ progenitor cells and examine their binding to the plant lectin Ulex europaeus I (Ulex). In normal bone marrow, 48.48 +/- 17.4% of the CD34+ cells bind to Ulex. Two color flow cytometry was used to sort CD34 + cells, and subsets of CD34+ cells, CD34+ Ulex+ and CD34+ Ulex-. These populations were sorted into colony assays to assess myeloid (CFU-GM) and erythroid (BFU-E) progenitors. The CD34+ Ulex+ subset was 84 +/- 14% BFU-E colonies (mean +/- S.D.) and had the highest cloning efficiency of 28 +/- 13%. Three color analysis of CD34+ Ulex+ cells showed staining with other erythroid (CD71, GlyA) antibodies and lack of stain. ing with myeloid (CD13, CD45RA) antibodies. These studies confirmed the erythroid characteristics of this subpopulation.
Antibodies↗
Acquired hypoprothrombinemia: effects of danazol treatment.
The lupus anticoagulant may be accompanied by an acquired factor II deficiency and bleeding. We report on a patient with a lupus anticoagulant and factor II (Fll) deficiency responsive to Danazol. Acquired hypoprothrombinemia (FII) with the lupus anticoagulant (LA) may be accompanied by a hemorrhagic diathesis. A 64-year-old male with discoid lupus erythematosis bled after an intestinal polypectomy. His FII level was 18%, and his FII antigen level was 20%. Danazol (D) (600 mg per day) administration was associated with a rise in FII activity and antigen to 50% within 10 days. The patient underwent abdominal surgery. We studied the effect(s) of D on the FII level and on other coagulation factors in this patient. The patient's plasma FII antigen had a single precipitin arc compared to the two peaks of normal plasma on counterimmunoelectrophoresis with Ca++. The samples pre- and during D therapy had the same positively charged arc as normal samples, although they were quantitatively different. Neuraminidase treatment demonstrated a decrease in the positively charged migration of normal and the patient's FII antigen. Affinity chromatography of normal and patient plasma on a Sepharose protein A column revealed FII antigen present in the patient's bound fraction. The relative percentages of bound FII before and during D treatment were similar. During D therapy, levels of FIX and X rose 50-100%, and protein C rose 20-25%, while free protein S did not change. D is an effective therapy for acquired FII deficiency associated with LA. D does not affect the binding of Ig to FII, but D raises FII levels by increasing synthesis of the FII protein.
Blood Loss, Surgical↗
Movement-related effects in fMRI time-series.
This paper concerns the spatial and intensity transformations that are required to adjust for the confounding effects of subject movement during functional MRI (fMRI) activation studies. An approach is presented that models, and removes, movement-related artifacts from fMRI time-series. This approach is predicated on the observation that movement-related effects are extant even after perfect realignment. Movement-related effects can be divided into those that are a function of position of the object in the frame of reference of the scanner and those that are due to movement in previous scans. This second component depends on the history of excitation experienced by spins in a small volume and consequent differences in local saturation. The spin excitation history thus will itself be a function of previous positions, suggesting an autoregression-moving average model for the effects of previous displacements on the current signal. A model is described as well as the adjustments for movement-related components that ensue. The empirical analyses suggest that (in extreme situations) over 90% of fMRI signal can be attributed to movement, and that this artifactual component can be successfully removed.
Artifacts↗
Late recurrences of vaginal clear cell adenocarcinoma.
Recurrent vaginal clear cell adenocarcinoma was diagnosed in two DES-exposed patients 17 and 19 years after initial therapy. These cases demonstrate the need for continued clinical evaluation, since patients with clear cell carcinoma of the vagina seem to be at greater risk for developing late recurrences than patients with squamous cell carcinomas.
Adenocarcinoma, Clear Cell↗
Transient coupling of Ng-CAM expression to NgCAM-dependent calcium signaling during migration of new neurons in the adult songbird brain.
The adult avian forebrain continues to generate neurons from subependymal zone (SZ) precursor cells, whose neuronal progeny migrate into the brain upon radial guide fibers. These neurons express the immunoglobulin-family adhesion molecule NgCAM, and their migration in culture is disrupted by anti-NgCAM Fab. Confocal imaging of adult zebra finch SZ loaded with the calcium indicator fluo-3, as well as ratio imaging with the indicator fura-2, revealed that migrating new neurons responded to microgram amounts of NgCAM with reversible increments in cytosolic calcium. The calcium response to NgCAM antigen was developmentally restricted, in that it was only manifested by neurons for roughly the 3- to 4-day period between 6 and 9 DIV, even though NgCAM expression persisted tonically thereafter. The period during which NgCAM elicited a calcium signal corresponded to the postmitotic age at which new, bipolar neurons leave the adult SZ to enter the brain parenchyma in vivo. Accordingly, the calcium response to NgCAM was largely limited to morphologically bipolar cells. Anti-NgCAM IgG also evoked a neuronal calcium signal over the same restricted period that NgCAM protein exerted its effect. These findings suggest a dynamic coupling and uncoupling of calcium-dependent signal transduction pathways to a stably expressed surface adhesion molecule, whose function in a given neuron may therefore evolve with cellular maturation.
Animals↗
Small bowel transplantation. A life-saving option for selected patients with intestinal failure.
Thirty-seven patients were listed for small bowel transplantation; 16 were transplanted and 15 died while waiting for a donor. Cyclosporine (N = 6) or tacrolimus (N = 10) were used for immune suppression. Graft rejection rates were lower in the combined liver/small bowel grafts than the isolated intestinal transplants (1/7 vs 5/7; P < 0.01) All of the cyclosporine group have died; the median survival was 25.7 months with two patients living more than five years. The tacrolimus group had fewer infections and a shorter hospital stay. All but two are alive with a median survival of 13 months. Seven of eight long-term survivors are off intravenous feedings. We conclude that small bowel transplantation is a life-saving option for patients with intestinal failure who cannot be maintained on total parenteral nutrition.
Adolescent↗
Allium sativum-induced death of African trypanosomes.
The effect of Allium sativum (Liliacea) on trypanosome-infected mice was examined. At a dose of 5.0 mg/ml, the oily extract from the pulp completely suppressed the ability of the parasites to be infective in the host. Column chromatography of the extract gave four fractions: ethylacetate/methanol, ethylacetate/ethanol, benzene/methanol, and acetic acid/methanol. Among these fractions, the acetic acid/methanol fraction retained the trypanocidal features of the crude extract. It cured experimentally infected mice of trypanosomiasis in 4 days when given at a dose of 120 mg/kg per day. The extract also manifested inhibition of procyclic forms of Trypanosoma brucei brucei and phospholipases from T. congolense, T. b. brucei, T. vivax. The extract appears to be diallyl-disulfide (DAD) and may interfere with the parasites' synthesis of membrane lipids.
Animals↗
A characterisation of dark-field imaging of colloidal gold labels in a scanning transmission X-ray microscope.
While X-ray microscopes provide images of biological specimens for which the contrast is mainly due to the difference in the absorption of carbon and oxygen when X-rays transmitted through the specimen are detected, signals other than absorption can also be used to form images. Using the Stony Brook scanning transmission X-ray microscope at the National Synchrotron Light Source, high-angle dark-field images have been formed of cells labelled with colloidal gold, with and without silver enhancement. The high density of the colloidal gold particles, or the silver particles seeded by the gold, leads to a large scattering signal, and the fact that the particle diameters are comparable to the width of the microscope point spread function results in good localisation of the label with high contrast. The dark-field images can have a greater signal to noise ratio than bright-field images acquired with the same incident X-ray dose. The theory of dark-and bright-field imaging is reviewed. Theoretical calculations of scattering from gold and silver particles are presented and good agreement is found between these and experimental dark-field images of 30 nm diameter gold particles. The signal to noise ratios of experimental bright-and dark-field images are measured and found to be in agreement with theory. Images are presented of cells labelled by immunolabelling and in situ hybridisation.
Fibroblasts↗