Search PubMed⌕ Search

Biomedical subjects

A Woods

Publications and source records attributed to A Woods.

At least 73 records · Page 4Linked to original sources

An evaluation of the Behring Diagnostics Enzygnost Syphilis enzyme immunoassay.

The Enzygnost Syphilis (Behring Diagnostics, Marburg, Germany) was evaluated (i) using sera already characterised using the rapid plasma reagin test and the Treponema pallidum hemagglutination test (TPHA); (ii) prospectively; (iii) using sera which are recognised as causing "problems" with enzyme immunoassays. The Enzygnost Syphilis was found to have a reproducibility of 98.6% and a sensitivity of 99.5% using sera known to be TPHA-reactive. When tested prospectively, the false reactive rate was 3.2%. These results show that the assay performs sufficiently satisfactorily for its introduction as a screening assay for evidence of syphilis infection in our population.

False Positive Reactions↗

Assessment of intubating conditions in adults after induction with propofol and varying doses of remifentanil.

We have assessed intubating conditions in three groups of 60 ASA I or II patients after induction of anaesthesia with propofol 2 mg kg-1 and remifentanil 0.5, 1.0 or 2.0 micrograms kg-1. Tracheal intubation was graded according to ease of laryn-goscopy, position of the vocal cords, coughing, jaw relaxation and movement of the limbs. Intubation was successful in 80%, 90% and 100% of patients after remifentanil 0.5, 1.0 or 2.0 micrograms kg-1, respectively. Overall intubating conditions were regarded as acceptable in 20%, 50% and 80% of patients, respectively. All three groups had a decrease in arterial pressure after induction but there was no difference between groups. The decrease in arterial pressure was not regarded as clinically significant. Intubating conditions were best after induction with remifentanil 2 micrograms kg and propofol 2 mg kg-1.

Adult↗

Caring for terminally ill persons: comparative analysis of attitudes (thanatophobia) of practicing physicians, student nurses, and medical students.

To investigate why some caregivers desire to avoid patients with terminal illnesses, a thanatophobia scale assessing caregivers' uncomfortable feelings and sense of helplessness was developed and evaluated among practicing physicians and student nurses and medical students. As a group, student nurses scored lower on the thanatophobia scale than practicing physicians and medical students. Higher scores on intolerance to clinical uncertainty were associated with higher thanatophobia scores in all groups. Scores for depressed mood, decreased sensitivity, and Machiavellianism were statistically significant predictors in some groups. Higher thanatophobia scores also predicted lower scores for ratings of self-esteem in caring for elderly patients. Among senior medical students, those with lower thanatophobia scores tended to enter primary care residencies. Health professional educators may find this scale helpful both in pinpointing sources of caregivers' angst and useful in counseling students in management of terminally ill persons.

Adult↗

Serine phosphorylation of syndecan-2 proteoglycan cytoplasmic domain.

Protein kinase C (PKC) is involved in cell-matrix and cell-cell adhesion, and the cytoplasmic domain of syndecan-2 contains two serines (residues 197 and 198) which lie in a consensus sequence for phosphorylation by PKC. Other serine and threonine residues are present but not in a consensus sequence. We investigated phosphorylation of syndecan-2 cytoplasmic domain by PKC, using purified GST-syndecan-2 fusion proteins and synthetic peptides corresponding to regions of the cytoplasmic domain. A synthetic peptide encompassing the entire cytoplasmic domain of syndecan-2 was phosphorylated by PKC with high affinity. Peptide mapping and substitution studies showed that both serines were phosphoacceptors, but each had slightly different affinity, with that of serine-197 being higher than serine-198. The efficiency of phosphorylation was concentration-dependent. At low concentrations, the cytoplasmic domain peptides were monomeric, with 2 mol/mol serine phosphorylation. At higher concentrations, however, the peptides formed dimers, with only 0.5 mol/mol phosphorylation. Concentration-dependent dimerization was not altered by phosphorylation. Phosphorylation is, therefore, dependent on the conformation of syndecan-2 cytoplasmic domain, but does not affect its oligomeric status.

Amino Acid Sequence↗

Multimerization of the cytoplasmic domain of syndecan-4 is required for its ability to activate protein kinase C.

The transmembrane proteoglycan syndecan-4, which is a coreceptor with integrins in cytoskeleton-matrix interactions, appears to be multimerized in vivo. Both purified and recombinant core proteins form sodium dodecyl sulfate-resistant oligomers, and we now report that a synthetic peptide corresponding to the central region of syndecan-4 cytoplasmic domain (4V) also oligomerizes. The degree of oligomerization correlates with the previously reported ability to bind protein kinase C (PKC) and regulate its activity. Only multimeric recombinant syndecan-4 core protein, but not the monomeric protein, potentiated the activity of PKCalpha, and only oligomeric syndecan-4 cytoplasmic peptides were active. Changes in peptide sequence caused parallel loss of stable oligomeric status and ability to regulate a mixture of PKCalphabetagamma activity. A synthetic peptide encompassing the whole cytoplasmic domain of syndecan-4 (4L) containing a membrane-proximal basic sequence did not form higher order oligomers and could not regulate the activity of PKCalphabetagamma unless induced to aggregate by phosphatidylinositol 4,5-bisphosphate. Oligomerization and PKC regulatory activity of the 4V peptide were both increased by addition of N-terminal cysteine and reduced by phosphorylation of the cysteine thiol group. Concentration of syndecan-4 at sites of focal adhesion formation may enhance multimerization and both localize PKC and potentiate its activity to induce stable complex formation.

Adenosine Triphosphate↗

Syndecan-4 proteoglycan regulates the distribution and activity of protein kinase C.

During cell-matrix adhesion, both tyrosine and serine/threonine kinases are activated. Integrin ligation correlates with tyrosine phosphorylation, whereas the later stages of spreading and focal adhesion and stress fiber formation in primary fibroblasts requires interactions of cell surface proteoglycan with heparin-binding moieties. This correlates with protein kinase C (PKC) activation, and PKCalpha can become localized to focal adhesions in normal, but not transformed, cells. PKC activation has been thought to be downstream of initial receptor-ligand interactions. We now show, however, that syndecan-4 transmembrane heparan sulfate proteoglycan and PKC co-immunoprecipitate and co-patch in vivo. The core protein of syndecan-4 can directly bind the catalytic domain of PKCalpha and potentiate its activation by phospholipid mediators. It can also directly activate PKCalpha in the absence of other mediators. This activity resides in the sequence LGKKPIYKK in the center of the short cytoplasmic domain, and other syndecans lack this sequence and PKC regulatory properties. Syndecan-4 is a focal adhesion component, and this interaction may both localize PKC and amplify its activity at sites of forming adhesions. This represents the first report of direct transmembrane signaling through cell surface proteoglycans.

Amino Acid Sequence↗

An HLA-DR1 transgene confers susceptibility to collagen-induced arthritis elicited with human type II collagen.

Rheumatoid arthritis (RA) is an autoimmune disease that is strongly associated with the expression of several HLA-DR haplotypes, including DR1 (DRB1*0101). Although the antigen that initiates RA remains elusive, it has been shown that many patients have autoimmunity directed to type II collagen (CII). To test the hypothesis that HLA-DR1 is capable of mediating an immune response to CII, we have generated transgenic mice expressing chimeric (human/mouse) HLA-DR1. When the DR1 transgenic mice were immunized with human CII (hCII), they developed a severe autoimmune arthritis, evidenced by severe swelling and erythema of the limbs and marked inflammation and erosion of articular joints. The development of the autoimmune arthritis was accompanied by strong DR1-restricted T and B cell responses to hCII. The T cell response was focused on a dominant determinant contained within CII(259-273) from which an eight amino acid core was defined. The B cell response was characterized by high titers of antibody specific for hCII, and a high degree of cross-reactivity with murine type II collagen. These data demonstrate that HLA-DR1 is capable of presenting peptides derived from hCII, and suggest that this DR1 transgenic model will be useful in the development of DR1-specific therapies for RA.

Adjuvants, Immunologic↗

Hydrogen peroxide-mediated alteration of the heme prosthetic group of metmyoglobin to an iron chlorin product: evidence for a novel oxidative pathway.

Treatment of metmyoglobin with H2O2 is known to lead to the crosslinking of an active site tyrosine residue to the heme [Catalano, C. E., Y. S. Choe, and P. R. Ortiz de Montellano (1989) J. Biol. Chem. 264, 10534-10541]. We have found in this study that this reaction also leads to an altered heme product not covalently bound to the protein. This product was characterized by visible absorption, infrared absorption, and mass and NMR spectrometry as an iron chlorin product formed from the saturation of the double bond between carbon atoms at positions 17 and 18 of pyrrole ring D with concomitant addition of a hydroxyl group on the carbon atom at position 18 and lactonization of the propionic acid to the carbon atom at position 17. Studies with the use of (18)O-labeled H2O2, O2, and H2O clearly indicate that the source of the added oxygen on the heme is water. Evidently, water adds regiospecifically to a cationic site formed on a carbon atom at position 18 after oxidation of the ferric heme prosthetic group with peroxide. Prolonged incubation of the reaction mixture containing the iron hydroxychlorin product led to the formation of an iron dihydroxychlorin product, presumably from a slow addition of water to the initial iron hydroxychlorin. The iron chlorin products characterized in this study are distinct from the meso-oxyheme species, which is thought to be formed during peroxide-mediated degradation of metmyoglobin, cytochrome P450, ferric heme, and model ferric hemes, and give further insight into the mechanism of H2O2-induced heme alterations.

Animals↗

The alpha1 and alpha2 isoforms of the AMP-activated protein kinase have similar activities in rat liver but exhibit differences in substrate specificity in vitro.

The AMP-activated protein kinase (AMPK) is a heterotrimeric complex composed of a catalytic subunit (alpha) and two regulatory subunits (beta and gamma). Two isoforms of the catalytic subunit (alpha1 and alpha2) have been identified. We show here that the alpha1- and alpha2-containing complexes contribute approximately equally to total AMPK activity in rat liver. Furthermore, expression of alpha1 or alpha2 with beta and gamma in mammalian cells demonstrates that both complexes have equal specific activity measured with the SAMS peptide. Using variant peptides, however, we show that alpha1 and alpha2 exhibit slightly different substrate preferences, which suggest that the two isoforms could play different physiological roles within the cell.

AMP-Activated Protein Kinases↗

Characterization of the AMP-activated protein kinase kinase from rat liver and identification of threonine 172 as the major site at which it phosphorylates AMP-activated protein kinase.

We have developed a sensitive assay for the AMP-activated protein kinase kinase, the upstream component in the AMP-activated protein kinase cascade. Phosphorylation and activation of the downstream kinase by the upstream kinase absolutely requires AMP and is antagonized by high (millimolar) concentrations of ATP. We have purified the upstream kinase >1000-fold from rat liver; a variety of evidence indicates that the catalytic subunit may be a polypeptide of 58 kDa. The physical properties of the downstream and upstream kinases, e.g. catalytic subunit masses (63 versus 58 kDa) and native molecular masses (190 versus 195 kDa), are very similar. However, unlike the downstream kinase, the upstream kinase is not inactivated by protein phosphatases. The upstream kinase phosphorylates the downstream kinase at a single major site on the alpha subunit, i.e. threonine 172, which lies in the "activation segment" between the DFG and APE motifs. This site aligns with activating phosphorylation sites on many other protein kinases, including Thr177 on calmodulin-dependent protein kinase I. As well as suggesting a mechanism of activation of AMP-activated protein kinase, this finding is consistent with our recent report that the AMP-activated protein kinase kinase can slowly phosphorylate and activate calmodulin-dependent protein kinase I, at least in vitro (Hawley, S. A., Selbert, M. A., Goldstein, E. G., Edelman, A. M., Carling, D., and Hardie, D. G. (1995) J. Biol. Chem. 270, 27186-27191).

AMP-Activated Protein Kinase Kinases↗

Syndecans, signaling, and cell adhesion.

Syndecans are transmembrane proteoglycans which can participate in diverse cell surface interactions, involving extracellular matrix macromolecules, growth factors, protease inhibitors, and even viral entry. Currently, all extracellular interactions are believed to be mediated by distinct structures within the heparan sulfate chains, leaving the roles of chondroitin sulfate chains and extracellular portion of the core proteins to be elucidated. Evidence that syndecans are a class of receptor involved in cell adhesion is mounting, and their small cytoplasmic domains may link with the microfilament cytoskeleton, thereby mediating signaling events. The molecular details are unknown, but the conservation of regions of syndecan cytoplasmic domains, and a strong tendency for homotypic association, support the idea that the ligand-induced clustering may be a discrete source of specific transmembrane signaling from matrix to cytoskeleton, as proposed for other classes of adhesion receptors.

Amino Acid Sequence↗

Characterization of AMP-activated protein kinase beta and gamma subunits. Assembly of the heterotrimeric complex in vitro.

There is growing evidence that mammalian AMP-activated protein kinase (AMPK) plays a role in protecting cells from stresses that cause ATP depletion by switching off ATP-consuming biosynthetic pathways. The active form of AMPK from rat liver exists as a heterotrimeric complex and we have previously shown that the catalytic subunit is structurally and functionally related to the SNF1 protein kinase from Saccharomyces cerevisiae. Here we describe the isolation and characterization of the two other polypeptides, termed AMPKbeta and AMPKgamma, that together with the catalytic subunit (AMPKalpha) form the active kinase complex in mammalian liver. Sequence analysis of cDNA clones encoding these subunits reveals that they are related to yeast proteins that interact with SNF1, providing further evidence that the regulation and function of AMPK and SNF1 have been conserved throughout evolution. The amino acid sequence of the beta subunit is most closely related to SIP2 (35% identity), while the amino acid sequence of the gamma subunit is 35% identical with SNF4. We show that both AMPKbeta and AMPKgamma mRNA and protein are expressed widely in rat tissues. We show that AMPKbeta interacts with both AMPKalpha and AMPKgamma in vitro, whereas AMPKalpha does not interact with AMPKgamma under the same conditions. These results suggest that AMPKbeta mediates the association of the heterotrimeric AMPK complex in vitro, and will facilitate future studies aimed at investigating the regulation of AMPK in vivo.

AMP-Activated Protein Kinases↗

Long-term treatment with lipoteichoic acid from Streptococcus faecalis affects differentiation and expression and cellular distribution of beta 1 integrins in human urothelial cells.

Gram-positive bacteria are recognized pathogens in urinary tract infections. Cellular mechanisms triggered by lipoteichoic acids (LTs), cell well components of gram-positive bacteria, have not been completely defined. We have postulated that infection-induced altered function of progenitors of urothelial cells residing in the basal layer is likely to have long lasting effects on the architecture and function of the urothelium. Our recent studies in vitro showed that treatment of poorly differentiated urothelial cells of basal type with LT from Streptococcus faecalis (LT-2) stimulated rapid proliferation of a subpopulation of progenitors of urothelial cells, supporting this possibility (Elgavish et al., 1996, J. Cell. Physiol., 169:42-51). The hypothesis underlying the present studies was that, following LT-triggered increase in proliferation of progenitors, the rate of differentiation of the resulting progeny was also stimulated. We proposed that this mechanism may allow rapid removal of cells from the injured area and replacement by cells that have not been exposed to infection. To simulate in vitro conditions in the basal layer that inhibit terminal differentiation, cells grew on fibronectin or collagen-coated substrate, in medium containing low Ca2+ (0.2 mM) and low levels of growth factors (0.005% bovine pituitary extract [BPE]). During the last 3 days in culture, cells grew in the same low Ca2+ (0.2 mM) medium, but without BPE, with or without LT-2. In a positive control group, cells grew during their last 3 days in culture in medium without BPE and LT-2 but in which levels levels of Ca2+ were higher (2 mM), a condition known to stimulate differentiation in other cell types. Several lines of evidence supported the possibility that long-term treatment with LT-2 stimulated progression of large colonies (i.e., the progeny resulting from LT-triggered proliferation) to a more differentiated state: (1) the rate of their differentiation, determined by criterion of intense cytokeratin 8 expression, was increased; (2) steady-state level of beta mRNA and expression of beta 1 subunit of integrins at the protein level were inhibited; (3) in contrast to large colonies in control cultures, the entire population of LT-2-treated large colonies contained beta 1 integrins distributed at cell-cell contacts. Raising extracellular Ca2+ concentration to 2 mM induced similar effects, suggesting that LT-2 may act by stimulating an increase in intracellular levels of Ca2+. However, further studies will be needed to elucidate the molecular mechanisms underlying the stimulatory effect of LT-2 on proliferation of progenitors of urothelial cells in the basal layer of the urothelium and subsequent differentiation of their progeny. We propose that these processes may have a causative role in the pathological changes that occur in the aftermath of chronic or recurrent suburothelial infection in the urinary bladder.

Cell Differentiation↗

Ambivalence and dialysis discontinuation.

Although dialysis discontinuation is the second leading "cause" of death among individuals who are maintained with chronic dialysis, little attention is accorded in the literature to patient ambivalence with this crucial decision. Three cases are presented of clinical situations involving termination of dialysis and ambivalence. The ambivalence of the patients is mirrored by the literature's vacillation between viewing cessation of life-support as pathological, and perceiving it as rational. There is considerable variability in the presentation of these cases, and they require exquisite sensitivity and flexibility on the part of medical personnel. As patient-physician communication is encouraged, and staff attempt to honor requests for initiation and/or discontinuation of treatment, we will see increasing numbers of ambivalent patients.

Adult↗

Experiences of battered women in health care settings: a qualitative study.

Interventions to help battered women seek and receive optimal health care must be informed by battered women's experiences in health care settings. In this study, we used a Systems Model to categorize the barriers battered women encounter in health care settings into patient, provider, and organizational levels. We conducted in-depth, face-to-face interviews with 31 battered women recruited by random digit dialing of households and by a publicity recruitment campaign. The data revealed that at the patient level, many women chose to conceal their abuse from their health care professionals, some fearing retaliation from their partners if they revealed the source of their injuries. At the provider level, the women perceived health care professionals to be disinterested or unsympathetic toward the needs of battered women, causing the women to feel ignored or trivialized. And at an organizational level, battered women believed that the structure of the health care system did not allow health care professionals enough time to deal with issues beyond treating their immediate presenting injuries. To ensure that battered women seek and receive optimal health care, multicomponent interventions should be designed to address the complex barriers at the three levels. We conclude by suggesting possible ways to help battered women get the tools they need to raise the issue of domestic violence with their health care professional. We also suggest ways to enable these professionals to identify battered women, validate their experiences, and provide appropriate referrals.

Adult↗

Self-esteem and caregivers' attitudes toward elderly persons.

A scale identifying 141 medical students who responded positively to geriatric patients was based on Rosenberg's Self-esteem Scale modified by adding a phrase about geriatric care. Personal and professional role traits that predicted a positive therapeutic attitude were high scores on social desirability or self-monitoring and low scores on thanatophobia and depression. Senior medical students who expressed the highest self-esteem toward caring for elderly people indicated family medicine as their first choice of residency.

Adult↗