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Biomedical subjects

A Gray

Publications and source records attributed to A Gray.

At least 73 records · Page 4Linked to original sources
PubMed2000

Analysis of the cellular functions of PTEN using catalytic domain and C-terminal mutations: differential effects of C-terminal deletion on signalling pathways downstream of phosphoinositide 3-kinase.

The tumour suppressor protein, PTEN (phosphatase and tensin homolog deleted on chromosome 10), is a phosphatase that can dephosphorylate tyrosine-containing peptides, Shc, focal adhesion kinase and phosphoinositide substrates. In cellular assays, PTEN has been shown to antagonize the PI-3K-dependent activation of protein kinase B (PKB) and to inhibit cell spreading and motility. It is currently unclear, however, whether PTEN accomplishes these effects through its lipid- or protein-phosphatase activity, although strong evidence has demonstrated the importance of the latter for tumour suppression by PTEN. By using a PTEN G129E (Gly(129)-->Glu) mutant that has lost its lipid phosphatase activity, while retaining protein phosphatase activity, we demonstrated a requirement for the lipid phosphatase activity of PTEN in the regulation of PKB activity, cell viability and membrane ruffling. We also made a small C-terminal deletion of PTEN, removing a putative PDZ (PSD95, Dlg and ZO1)-binding motif, with no detectable effect on the phosphatase activity of the protein expressed in HEK293 cells (human embryonic kidney 293 cells) assayed in vitro. Surprisingly, expression of this mutant revealed differential requirements for the C-terminus in the different functional assays. Wild-type and C-terminally deleted PTEN appeared to be equally active in down-regulating PKB activity, but this mutant enzyme had no effect on platelet-derived growth factor (PDGF)-induced membrane ruffling and was only partially active in a cell viability assay. These results stress the importance of the lipid phosphatase activity of PTEN in the regulation of several signalling pathways. They also identify a mutation, similar to mutations that occur in some human tumours, which removes the effect of PTEN on membrane ruffling but not that on PKB.

Base Sequence↗
PubMed2000

Centre-specific or average unit costs in multi-centre studies? Some theory and simulation.

Costing issues are increasingly being addressed in multi-centre studies. In this paper, two methods for collecting costing information are compared within a simulated clinical trial setting. One method estimates average treatment costs by applying unit costs averaged across treatment centres to centre-specific volumes of resource use. The second uses centre-specific information for both the unit costs and the resource volumes, and then averages across centres. Using a pre-specified production relation between the different volumes of resource use, and simulating changes in unit costs, it is shown that these two methods result in statistically different estimates of average treatment costs. This finding holds, regardless of the degree of substitutability between the resource volumes, except when considerable uncertainty surrounds treatment centre responses to relative changes in unit costs. The findings suggest that a more cautious approach should be adopted in the collection, calculation and interpretation of treatment costs in multi-centre studies.

Computer Simulation↗
PubMed2000

Case management for people with severe mental disorders.

BACKGROUND: Since the 1960s, in many parts of the world, large psychiatric were closed down and people were treated in outpatient clinics, day centres or community mental health centres. Rising readmission rates suggested that this type of community care may be less effective than anticipated. In the 1970s case management arose as a means of co-ordinating the care of severely mentally ill people in the community. OBJECTIVES: To determine the effects of case management as an approach to caring for severely mentally ill people in the community. Case management was compared against standard care on four main indices: (i) numbers remaining in contact with the psychiatric services; (ii) extent of psychiatric hospital admissions; (iii) clinical and social outcome; and (iv) costs. SEARCH STRATEGY: Electronic searches of CINAHL (1997), the Cochrane Schizophrenia Group's Register of trials (1997), EMBASE (1980-1995), MEDLINE (1966-1995), PsycLIT (1974-1995) and SCISEARCH (1997) were undertaken. References of all identified studies were searched for further trial citations. SELECTION CRITERIA: The inclusion criteria were that studies should be randomised controlled trials that (i) had compared case management to standard community care; and (ii) had involved people with severe mental disorder mainly between the ages of 18-65. Studies of case management were defined as those in which the investigators described the intervention as 'case' or 'care' management rather than 'Assertive Community Treatment' or 'ACT'. DATA COLLECTION AND ANALYSIS: A study was carried out to test the reliability of the inclusion criteria. Categorical data were extracted twice and then cross-checked, any disagreements being resolved by discussion. Odds ratios and the number needed to treat were estimated. Continuous data collected by a measuring instrument was only included if the instrument (i) had been described in a peer-reviewed journal; (ii) was a self-report or had been completed by an independent rater; and (iii) provided a summary score for a broad area of functioning. Normally distributed continuous data were included if means and standard deviations were available. Non-normal data were included if analysed either after transformation or using non-parametric methods. Tests for heterogeneity were conducted. MAIN RESULTS: Case management increased the numbers remaining in contact with services (for case management odds ratio = 0.70; 99%CI 0.50-0. 98; n=1210). Case management approximately doubled the numbers admitted to psychiatric hospital (OR 1.84; 99% CI 1.33-2.57; n=1300). Except for a positive finding on compliance, from one study, case management showed no significant advantages over standard care on any psychiatric or social variable. Cost data did not favour case management but insufficient information was available to permit definitive conclusions. REVIEWER'S CONCLUSIONS: Case management ensures that more people remain in contact with psychiatric services (one extra person remains in contact for every 15 people who receive case management), but it also increases hospital admission rates. Present evidence suggests that case management also increases duration of hospital admissions, but this is not certain. Whilst there is some evidence that case management improves compliance, it does not produce clinically significant improvement in mental state, social functioning, or quality of life. There is no evidence that case management improves outcome on any other clinical or social variables. Present evidence suggests that case management increases health care costs, perhaps substantially, although this is not certain. In summary, therefore, case management is an intervention of questionable value, to the extent that it is doubtful whether it should be offered by community psychiatric services. It is hard to see how policy makers who subscribe to an evidence-based approach can justify retaining case management as 'the cornerstone' of community mental hea

Case Management↗
PubMed2000

Life-expectancy projection by modelling and computer simulation (UKPDS 46).

A method is described for estimating the life-expectancy of cohorts of type 2 diabetic patients, based on computer simulation from a parametric model. The method can be used where non-parametric methods, such as the Kaplan-Meier estimate, fail due to lack of the data. The simulation algorithm combines observed and modelled information to estimate the life-expectancy implications of event rates observed within a study. The use of bootstrap methods to estimate confidence intervals is discussed. The methods are illustrated with results that have been previously published, without derivation, in a health economic analysis of tight blood pressure (BP) control in the UK Prospective Diabetes Study (UKPDS), and the application to other health economic analyses of UKPDS data is discussed.

Algorithms↗
PubMed2000

Characterization of huJAM: evidence for involvement in cell-cell contact and tight junction regulation.

Cell-cell interactions of the mucosal epithelia are important for the maintenance and establishment of epithelial barrier function. During events of inflammation, such cell-cell interactions are often disrupted, resulting in a leaky epithelial barrier, which in turn can lead to various inflammatory and infective dysfunctions. Human junctional adhesion molecule (huJAM), found on the mucosal epithelia and vascular endothelia of many major organ systems, is a membrane glycoprotein which resolves to a doublet band of approximately 40 and approximately 37 kDa under SDS-PAGE analysis, representing differentially glycosylated forms of the same protein. huJAM was localized to the lateral membrane of Caco-2 cells (a human colonic epithelial cell line) monolayers, in an area basolateral of the epithelial tight junctions (TJ). Through functional and biochemical assays, we show huJAM to be able to homotypically associate and to participate in TJ restitution after trypsin-EDTA disruption. Furthermore, we also observed a migration of huJAM expression toward areas of cell-cell contacts during events of cell adhesion and monolayer formation. These qualities makes huJAM a likely player in the regulation of cell-cell contacts and the subsequent formation of TJs.

Animals↗
PubMed2000

Health economics and orthopaedics.

It is becoming widely accepted that research which considers only the outcome and not the costs associated with new technologies in health care, is of limited value in making decisions about the use of scarce resources. Economic evaluation is becoming a standard feature of clinical research but many published economic evaluations fall short of best practice in their methodology. We have described the essential features of economic evaluation, using published studies in orthopaedics, in order to try to improve the ability of orthopaedic surgeons to read, understand and appraise such studies critically, and to encourage them to consider including economic evaluation in future investigations.

Cost-Benefit Analysis↗
PubMed2000

United Kingdom national confidential enquiry into perioperative deaths.

The enquiry (NCEPOD) was established to examine simultaneously the anaesthetic and surgical circumstances surrounding the death of a patient undergoing surgery, it reviewed clinical practice and identified remedial factors in the practice of anaesthesia and surgery in order to improve the quality of care. Data were reported on every death occurred within 30 days of a procedure performed by a surgeon or gynaecologist under general or local anaesthesia, excepting obstetric deaths, and were collected int he sample of questionnaires to surgeons and anaesthesists involved. In high risk patient the optimisation of cardiovascular system with fluids, inotropic agents, beta-blockers and invasive monitoring may improve mortality (a speculative calculation reported 1,700 out 19,000 lives saved in 1994/1995 in UK). The majority of deaths occurred in emergency operations, urgent operations delayed for no operating theatre free during the day, training personnel which performed emergency operations during the night and/or postoperative care provided by non specialized clinicians, hospital not equipped with high dependency units (HDU), surgery undertaken although the predictable infaust outcome, they were all problems outlooked by the investigation. In conclusion improvements are necessary in hospital facilities and working pattern of surgeons and anaesthesists.

Humans↗
PubMed1999

The pleckstrin homology domains of protein kinase B and GRP1 (general receptor for phosphoinositides-1) are sensitive and selective probes for the cellular detection of phosphatidylinositol 3,4-bisphosphate and/or phosphatidylinositol 3,4,5-trisphosphate in vivo.

We have tested the binding specificities of the pleckstrin homology (PH) domains of protein kinase B (PKB) and GRP1 (general receptor for phosphoinositides-1), expressed as green fluorescent protein (GFP) fusion proteins [PH(PKB)GFP and PH(GRP1)GFP respectively] in HEK 293 cells and Swiss 3T3 cells, using confocal microscopy. Stimulation of HEK 293 cells with insulin caused a small, but sustained, increase in PtdIns(3,4,5)P(3) levels, detected using a radioligand displacement assay, which was mirrored by the translocation of PH(PKB)GFP and PH(GRP1)GFP from the cytosol to the plasma membrane of live, transfected cells. Similar results were obtained using Swiss 3T3 cells stimulated with platelet-derived growth factor (PDGF) and expressing either PH(PKB)GFP or PH(GRP1)GFP. Biochemical analyses confirmed the accumulation of both PtdIns(3,4,5)P(3) and PtdIns(3,4)P(2) in response to PDGF, but only the latter was present at increased levels in Swiss 3T3 cells 30 min after an oxidative stress (1 mM H(2)O(2)). Concomitantly, only PH(PKB)GFP, and not PH(GRP1)GFP, was localized at plasma membranes after 30 min of treatment with H(2)O(2). The fusion proteins appear accurately to report the spatial and temporal distribution of PtdIns(3,4,5)P(3) and PtdIns(3,4)P(2) in intact cells. These results establish the lipid selectivity of these PH domains in vivo, and further emphasize the overlapping, but distinct, second messenger roles of PtdIns(3,4,5)P(3) and PtdIns(3,4)P(2).

Androstadienes↗
PubMed1999

Distinct phosphatidylinositol 3-kinase lipid products accumulate upon oxidative and osmotic stress and lead to different cellular responses.

Signaling by phosphatidylinositol (PI) 3-kinases is mediated by 3-phosphoinositides, which bind to Pleckstrin homology (PH) domains that are present in a wide spectrum of proteins. PH domains can be classified into three groups based on their different lipid binding specificities. Distinct 3-phosphoinositides can accumulate upon PI 3-kinase activation in cells in response to different stimuli and mediate specific cellular responses. In Swiss 3T3 mouse fibroblasts, oxidative stress induced by 1 mM H(2)O(2) caused almost exclusive accumulation of phosphatidylinositol 3,4-bisphosphate (PtdIns(3, 4)P(2)), whereas osmotic stress increased both phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P(3)) and PtdIns(3,4)P(2) levels. The increase in PtdIns(3,4)P(2) levels, caused by oxidative stress, correlated with the activation of protein kinase B, which has a promiscuous PH domain that binds both PtdIns(3,4,5)P(3) and PtdIns(3, 4)P(2). p70 S6 kinase, another signaling component downstream of PI 3-kinase, however, was not activated by this oxidative stress-induced increase in PtdIns(3,4)P(2) levels. Increased PtdIns(3,4,5)P(3) and PtdIns(3,4)P(2) levels in response to osmotic stress did not correlate with protein kinase B activation, because of concomitant activation of an inhibitory pathway, but p70 S6 kinase was activated by osmotic stress. These results demonstrate that PtdIns(3,4)P(2) can accumulate independently of PtdIns(3,4, 5)P(3) and exerts a pattern of cellular responses that is distinct from that induced by accumulation of PtdIns(3,4,5)P(3).

3T3 Cells↗
PubMed1999

A role for AMPA/kainate receptors in conditioned place preference induced by diazepam in the rat.

There is increasing evidence for a role of glutamate receptors in the reinforcing properties of dependence producing drugs such as the psychostimulants and opiates. Activation of AMPA/kainate receptors are implicated in the acquisition of amphetamine-induced reinforcement but a role for this receptor in benzodiazepine-induced reinforcement has not been examined. In the present study the ability of the orally active AMPA/kainate antagonist GYKI 52466 was assessed for its ability to block the reinforcing properties of diazepam in a conditioned place preference paradigm. Diazepam (2.5 and 5.0 mg/kg, i.p.) produced a robust place preference and GYKI 52466 inhibited the acquisition of place preference conditioning-induced by diazepam. These results suggest that glutamatergic pathways are an important component of the circuitry involved in the acquisition of a benzodiazepine induced place preference.

Animals↗
PubMed1999

The mechanism of inhibition of beta-oxidation by aspirin metabolites in skin fibroblasts from Reye's syndrome patients and controls.

The effects of aspirin metabolites on beta-oxidation were studied in skin fibroblasts from eight typical Reye's syndrome (RS) patients and controls. RS patients' cells did not differ from controls in rates of palmitate oxidation or in the three component activities of the mitochondrial trifunctional enzyme (MTE), indicating no inherited beta-oxidation defect. Aspirin metabolites salicylate, hydroxyhippurate and gentisate, but not aspirin, directly inhibited palmitate oxidation in control and RS cells. RS cells were significantly more sensitive to inhibition than controls at 0.5 to 5 mM salicylate. Inhibition was concentration-dependent and reversible. Inhibition did not occur in fibroblasts lacking activity of the long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) activity of MTE. Salicylate was therefore inhibiting beta-oxidation at this step. Hydroxyhippurate and salicylate reversibly inhibited HAD activities in extracts of control and RS cells. Studies with pure short-chain HAD and LCHAD (MTE) showed hydroxyhippurate and salicylate were competitive inhibitors of the former but mixed (not competitive) inhibitors of the latter. Both compounds inhibited the combined, three-step, MTE reaction measured in the physiological direction. We conclude that (1) salicylate and hydroxyhippurate decrease beta-oxidation in intact cells by reversible inhibition of LCHAD activity of the MTE, and (2) beta-oxidation in RS cells is inherently more sensitive to inhibition by low concentrations of salicylate than controls.

3-Hydroxyacyl CoA Dehydrogenases↗
PubMed1999

Role of phosphatidylinositol 3,4,5-trisphosphate in regulating the activity and localization of 3-phosphoinositide-dependent protein kinase-1.

3-Phosphoinositide-dependent protein kinase-1 (PDK1) interacts stereoselectively with the d-enantiomer of PtdIns(3,4,5)P3 (KD 1.6 nM) and PtdIns(3,4)P2 (KD 5.2 nM), but binds with lower affinity to PtdIns3P or PtdIns(4,5)P2. The binding of PtdIns(3,4,5)P3 to PDK1 was greatly decreased by making specific mutations in the pleckstrin homology (PH) domain of PDK1 or by deleting it. The same mutations also greatly decreased the rate at which PDK1 activated protein kinase Balpha (PKBalpha) in vitro in the presence of lipid vesicles containing PtdIns(3,4,5)P3, but did not affect the rate at which PDK1 activated a PKBalpha mutant lacking the PH domain in the absence of PtdIns(3,4,5)P3. When overexpressed in 293 or PAE cells, PDK1 was located at the plasma membrane and in the cytosol, but was excluded from the nucleus. Mutations that disrupted the interaction of PtdIns(3,4,5)P3 or PtdIns(4,5)P2 with PDK1 abolished the association of PDK1 with the plasma membrane. Growth-factor stimulation promoted the translocation of transfected PKBalpha to the plasma membrane, but had no effect on the subcellular distribution of PDK1 as judged by immunoelectron microscopy of fixed cells. This conclusion was also supported by confocal microscopy of green fluorescent protein-PDK1 in live cells. These results, together with previous observations, indicate that PtdIns(3,4,5)P3 plays several roles in the PDK1-induced activation of PKBalpha. First, it binds to the PH domain of PKB, altering its conformation so that it can be activated by PDK1. Secondly, interaction with PtdIns(3,4,5)P3 recruits PKB to the plasma membrane with which PDK1 is localized constitutively by virtue of its much stronger interaction with PtdIns(3,4,5)P3 or PtdIns(4,5)P2. Thirdly, the interaction of PDK1 with PtdIns(3,4,5)P3 facilitates the rate at which it can activate PKB.

3-Phosphoinositide-Dependent Protein Kinases↗
PubMed1999

Isolation and Characterization of Twelve Microsatellite Loci for Rockfish (Sebastes).

: We describe the first microsatellites for rockfishes in the diverse genus Sebastes. Clones containing microsatellites were isolated from the genomic library of a quillback rockfish, Sebastes maliger. Twelve microsatellites are characterized; six of these are polymorphic in quillback rockfish, and eight are polymorphic in at least one rockfish species on which they were tested. The number of alleles per variable locus ranged from 4 to 15 and averaged 6.8. The expected heterozygosities ranged from 0.38 to 0.79 and averaged 0.60 in these loci. These loci should prove valuable in studies examining species identification, population genetics, hybridization, paternity, kinship, and microsatellite evolution.

Journal Article↗
PubMed1999

Developmental and etiological characteristics of children with sexual behavior problems: treatment implications.

OBJECTIVE: Baseline data are reported on the demographics, psychological adjustment, victimization, and perpetration histories of 127 6- to 12-year-old children who have engaged in developmentally unexpected sexual behaviors. Information regarding the children's caregivers, and their extended families, is also presented. Data were collected during intake of the families into a longitudinal treatment outcome study. METHOD: A comprehensive battery of psychometric devices and a structured interview were completed with 127 children with sexual behavior problems and their primary caregivers at intake to a treatment outcome study. RESULTS: More than half of the children engaging in developmentally unexpected sexual behaviors had been abused both sexually and physically by more than two different perpetrators. One-third of the people who had maltreated these children were less than 18 years old. These children had acted out against an average of two other children. High levels of distress in the children and their caregivers were evident across a number of psychometric and historical variables. CONCLUSION: Children with sexual behavior problems exhibited a number of functional impairments commonly associated with maltreatment, including learning and psychiatric disorders. Their caregivers and families manifested several characteristics that deter children's recovery from maltreatment, including an impaired attachment between parent and child. The scope of the children's problems requires that treatment extend beyond the therapist's office to include schools and other agencies or individuals with whom the child and families have regular contact.

Analysis of Variance↗
PubMed1999

Fibrinogen.

Fibrinogen is a blood-borne glycoprotein comprised of three pairs of nonidentical polypeptide chains. Following vascular injury, fibrinogen is cleaved by thrombin to form fibrin which is the most abundant component of blood clots. As well as controlling blood loss at sites of tissue damage, other properties of fibrinogen have recently been discovered. For example, various cleavage products of fibrinogen and fibrin, released during coagulation and fibrinolysis, respectively, regulate cell adhesion and spreading, display vasoconstrictor and chemotactic activities, and are mitogens for several cell types including fibroblasts, endothelial and smooth muscle cells. Current research aims to define the bioactive fibrinogen molecule moieties and cellular receptors involved in these processes. Future studies may provide us with new opportunities to develop agents which are useful in promoting tissue repair or conversely in inhibiting fibrosis in inflammatory and fibroproliferative diseases where endothelial cell damage or chronic leakage of blood proteins is a feature.

Cardiovascular Agents↗