Effect of retinal and/or extra-retinal information on age in memory-guided saccades.

The aim of the study was to determine whether the addition of retinal or extra-retinal information could be used to improve memory-guided saccadic performance in healthy participants. Furthermore, we included two age groups in our study; healthy young adult subjects (mean age 20 years) and healthy middle-aged adult subjects (mean age 52 years). All subjects performed a novel task that incorporated a Go/NoGo task with a memory-guided saccade paradigm to investigate whether extra-retinal information (making a saccade towards the visible target) or retinal (a visible frame-of-reference) has any affect on the accuracy or variability of the response. We found all subjects made slight hypometric responses to the memory-guided targets. Both younger and middle-aged subjects revealed an increase in accuracy in the Go task compared with the NoGo task and the framed condition compared to the frameless condition, respectively. The frame also revealed a significant decrease in variability in the memory-guided saccades. A positive correlation in errors between the 1st and 2nd saccade in the Go task was revealed for all subjects; however, the older subjects revealed a greater correlation than younger subjects. The results presented indicate that younger and middle-aged perform highly similar patterns of errors during eye movements to remembered locations. However, middle-aged subjects show a greater tendency to use extra-retinal and retinal feedback to guide the response.

Protease-resistant form of insulin-like growth factor-binding protein 5 is an inhibitor of insulin-like growth factor-I actions on porcine smooth muscle cells in culture.

IGFs are pleiotrophic mitogens for porcine smooth muscle cells (pSMC) in culture. The effects of IGFs on cells are modulated by various insulin-like growth factor-binding proteins (IGFBP). IGFBP-5 is synthesized by pSMC and binds to the extracellular matrix. However, IGFBP-5 is also secreted into conditioned medium of cultured cells and is cleaved into fragments by a concomitantly produced protease. These fragments have reduced affinity for the IGFs and cleavage makes it difficult to assess the role of intact IGFBP-5. To study the consequence of accumulation of intact IGFBP-5 in medium, we determined the cleavage site in IGFBP-5 and prepared a protease resistant mutant. Amino acid sequencing of purified IGFBP-5 fragments suggested Arg138-Arg139 as the primary cleavage site. Arg138-Arg139-->Asn138-Asn139 mutations were introduced to create protease-resistant IGFBP-5, which has the same affinity for IGF-I as the native protein. This mutant IGFBP-5 remained intact even after 24 h of incubation and it inhibited several IGF-I actions when added to pSMC culture medium. The mutant IGFBP-5 (500 ng/ml) decreased IGF-I stimulated cellular DNA synthesis by 84%, protein synthesis by 77%, and it inhibited IGF-I stimulated migration of pSMC by 77%. It also inhibited IGF-I stimulation of IRS-1 phosphorylation. In contrast, the same amount of native IGFBP-5 did not inhibit IGF-I actions. The significance of inhibitory effects of the protease resistant IGFBP-5 was further demonstrated in pSMC transfected with mutant or native IGFBP-5 cDNAs. The mutant IGFBP-5 accumulated in culture medium of transfected cells, while native IGFBP-5 was degraded into fragments, PSMC overexpressing the mutant IGFBP-5 also responded poorly to IGF-I compared with mock transfected cells. IGF-I (5 ng/ml) increased [35S]methionine incorporation into control cells by 36% above the basal level, but it did not significantly change (4%) in pSMC cultures that were producing the mutant IGFBP-5. In conclusion, the accumulation of protease-resistant IGFBP-5 in the medium was inhibitory to IGF-I actions on pSMC. This suggests that proteolysis can prevent IGFBP-5 from acting as an inhibitor of IGF-I-stimulated effects and that it serves as an important mechanism for regulating cellular responsiveness to IGF-I.

Insulin-like growth factor-binding protein-5 is cleaved by physiological concentrations of thrombin.

Insulin-like growth factor (IGF)-binding protein-5 (IGFBP-5) is cleaved by a serine protease that is secreted by fibroblasts and porcine smooth muscle cells (pSMC) in culture. To investigate whether other serine proteases could cleave this substrate at physiologically relevant concentrations, we determined the proteolytic effects of thrombin on IGFBP-5. Human alpha-thrombin (0.0008 NIH U/ml) cleaved IGFBP-5 into 24-, 23-, and 20-kDa non-IGF-I-binding fragments. Cleavage occurred at a physiologically relevant thrombin concentration. The effect was specific for IGFBP-5, as other forms of IGFBPs, e.g. IGFBP-1, IGFBP-2, and IGFBP-4 were not cleaved by thrombin. Although IGFBP-3 was cleaved by thrombin, this effect required a 50-fold greater thrombin concentration. [35S]Methionine labeling followed by immunoprecipitation confirmed that IGFBP-5 that was constitutively synthesized by pSMC cultures was also degraded by thrombin into 24-, 23-, and 20-kDa fragments. The binding of IGF-I to IGFBP-5 partially inhibited IGFBP-5 degradation by thrombin, and an IGF analog that does not bind to IGFBP-5 had no effect. Thrombin did not account for the serine protease activity that had been shown previously to be present in pSMC-conditioned medium. This was proven by showing that 1) no immunoreactive thrombin could be detected in the pSMC-conditioned medium; 2) the IGFBP-5 fragments that were generated by thrombin showed three cleavage sites (Arg192-Ala193, Arg156-Ile157, and Lys120-His121), whereas the serine protease in conditioned medium cleaves IGFBP-5 at a different site; and 3) hirudin had no effect on IGFBP-5 cleavage by the protease in pSMC medium; however, it inhibited IGFBP-5 degradation by thrombin. To determine the physiological significance of IGFBP-5 cleavage, the effect of an IGFBP-5 mutant that is resistant to cleavage by the pSMC protease and has been shown to inhibit IGF-I actions in pSMC was determined. This mutant inhibited IGF-I-stimulated DNA synthesis, but if thrombin was added simultaneously, IGF-I was fully active. In summary, physiological concentrations of thrombin degrade IGFBP-5. Degradation can be blocked by hirudin and is partially inhibited by IGF-I binding. Generation of active thrombin in vessel walls may be a physiologically relevant mechanism for controlling IGF-I bioactivity.

A protease-resistant form of insulin-like growth factor (IGF) binding protein 4 inhibits IGF-1 actions.

Smooth muscle cells (SMC) secrete a serine protease that cleaves insulin-like growth factor (IGF) binding protein (IGFBP)-4 into fragments that have low affinity for IGF-1. When IGFBP-4 is added to monolayer cultures of cell types that do not secrete this protease, IGF-1 stimulation of DNA synthesis is significantly inhibited. In contrast, if cell types that secrete this protease are used, IGFBP-4 is a much less potent inhibitor. These studies were conducted to determine whether proteolysis of IGFBP-4 accounted for its reduced capacity to inhibit IGF-1-stimulated DNA synthesis. The cleavage site in IGFBP-4 that the SMC protease uses was determined to be lysine120, histidine121. A protease-resistant mutant form of IGFBP-4 was prepared, expressed, purified, and tested for biologic activity using porcine SMC cultures. Addition of the protease-resistant mutant resulted in inhibition of DNA and cell migration responses to IGF-1. The inhibition was concentration dependent and was maximal when 500 ng/ml (20 nM) of the mutant was added with 20 ng/ml (2.8 nM) of IGF-1. When the mutant was added in the absence of IGF-1, it had no activity. The results show that cleavage of IGFBP-4 at lysine120, histidine121 results in inactivation of the ability of IGFBP-4 to bind to IGF-1. Creation of a mutant form of IGFBP-4 that was not cleaved by the protease resulted in inhibition of IGF-1-stimulated actions. The results suggest that IGFBP-4 can act as a potent inhibitor of the anabolic effects of IGF-1 and that the variables that regulate protease activity may indirectly regulate IGF-1 actions.

Complex mediation of uterine endometrial epithelial cell growth by insulin-like growth factor-II (IGF-II) and IGF-binding protein-2.

The coexpression of IGF (-I and -II) peptides, corresponding receptors, and IGF binding proteins (IGFBPs) in uterine endometrium suggests that a significant component of IGF action in this tissue is via autocrine or paracrine pathways, or both. The present study examined whether IGF-II and a major uterine-expressed IGF-II binding protein, IGFBP-2, modulate endometrial epithelial cell mitogenesis. Serum-deprived porcine endometrial glandular epithelial (GE) cells of early pregnancy were treated with various concentrations of IGFs, recombinant porcine (rp) IGFBP-2, or both, and examined for changes in cellular mitogenesis by incorporation of [(3)H]thymidine into DNA. Recombinant human (rh) IGF-II stimulated DNA synthesis in a dose-dependent manner. Human [Leu(27)]-IGF-II, an analog with selective affinity for the IGF-II (type II) receptor, increased thymidine uptake by twofold compared with untreated GE cells. When added in combination with an equimolar concentration of rhIGF-I, [Leu(27)]-IGF-II or rhIGF-II stimulated thymidine incorporation to a greater extent than did rhIGF-I alone. Ligand blot analysis of GE cell conditioned medium revealed the presence of four IGFBPs with molecular masses of 48, 31, 23, and 15 kDa. Physiological concentrations of rpIGFBP-2 (nM range) increased both basal and IGF-induced DNA synthesis in GE cells. At equimolar concentrations, Des(1-6)IGF-II (an IGF-II analog with much reduced affinity for IGFBPs) and rpIGFBP-2 had additive effects on GE cell mitogenesis, suggesting that the IGFBP-2 modulation of uterine cell growth may involve both IGF-dependent and IGF-independent pathways. Our results demonstrate the complex interplay of IGF system components in uterine endometrial epithelial growth regulation in vitro, identify IGF-II and IGFBP-2 as locally coexpressed uterine epithelial cell mitogens, and suggest the presence of a functional signaling pathway by which IGF-II stimulates epithelial cell proliferation via the type II receptor.

The mechanism of bioactivation and antigen formation of amodiaquine in the rat.

A glutathione conjugate of amodiaquine has been isolated and characterized from rat bile after administration of [14C]amodiaquine (50 mumol/kg, 5.0 muCi/rat) to anaesthetized male Wistar rats. Thioether conjugates of amodiaquine in rat bile accounted for a total of 12% of the dose, 5 hr after administration of the drug. In addition, 1% of the dose remained in the liver covalently bound to tissue proteins after 5 hr. These findings provide direct evidence that a chemically reactive metabolite, amodiaquine quinoneimine, has been formed from the drug in vivo. A second major metabolite, desethylamodiaquine, accounting for 14% of the given dose, was present in the liver after 5 hr. Enzyme inhibition studies with ketoconazole-pretreated rats showed that both amodiaquine quinoneimine and desethylamodiaquine formation can be catalysed by cytochrome P450. The demonstration that amodiaquine readily and extensively forms a metabolite in vivo, with strong reactivity towards protein and non-protein thiol groups, may help to explain the idiosyncratic toxicity observed in man.

An investigation into causes of resistance of a cloned line of BHK cells to a strain of foot-and-mouth disease virus.

The reduced ability of foot-and-mouth disease virus (FMDV) strain Asia 1 Iran 1/73 to replicate in the cloned BHK cell line AA7 was not due to lack of virus attachment at the cell surface. Instead, the main restriction in the viral growth cycle occurred during synthesis and processing of viral macromolecules, and/or during the earliest stages of their assembly. Reduced efficiency of penetration and uncoating of virus attached to the cells may also have contributed to inhibition of virus replication. Viral components or subviral particles did not accumulate and defective interfering particles were not detected. The reduced number of infective virions produced was released from infected cells at the normal rate. No interferon production could be demonstrated.

Chipping hammer vibration.

An investigation was carried out to determine the factors influencing the vibration of chipping hammers and to find ways of reducing the vibration at the operator's hand. It showed that substantially higher vibration levels are produced at the chisel of a chipping hammer than at its handle. Typical weighted values for the two areas were 24 and 8 m/s2, respectively. This finding agrees with medical observations showing that invariably the hand holding the chisel exhibits the more severe symptoms of vibration-induced white finger. A sleeve which fits onto the chisel was developed which can effect a reduction in vibration of up to 66%. A prototype hammer was developed incorporating an isolating material which reduced the weighted vibration value from 7 to 3 m/s2. Vibration isolating gloves were tested and resulted in an additional reduction in vibration of up to 63% when used with the chisel sleeve. A unique mounting device using rubber isolators was designed which protects the accelerometer from very intense high-frequency vibration but allows accurate measurement of chisel vibration in the frequency range of interest.

Hermeneutic analysis: a qualitative decision trail.

Qualitative analysis is problematic from two perspectives, which exist in the science and art debate [Tesch, R., 1990. Qualitative Research. Analysis types and software tools. The Falmer Press, London; Robson, C., 1993. Real World Research. Blackwell, Oxford]. The science view claims that the absence of clear and agreed analysis processes, which can be found in the quantitative domain, attracts labels to qualitative analysis of intuitive artistry and personal journeys which are considered 'unscientific'. This thinking remains dominant despite the growth of systematic qualitative analysis supported through computer analysis systems [Tesch, 1990]. However, in the art domain there is real resistance to the development of set methods of analysis which does view qualitative analysis, "as more of an art than a science" [Robson, 1993, p. 370]. This paper offers a contribution to the qualitative analysis tension through the promotion and illustration of a decision making trail. This option supports the principles of academic rigour in qualitative research [Guba, E.G., Lincoln, Y.S., 1981. Effective Evaluation. Jossey Bass, San Francisco] as a decision trail permits the research community to make their own judgements concerning the process of analysis, the overall trustworthiness of the research and therefore its presented interpretations.

Ethical issues in health education.

Moral dilemmas abound during health education practice and ethical decisions have to be made. This article examines the contribution of the four guiding ethical principles (respect for autonomy, beneficence, non-maleficence and justice) to nurse decision making. Multidisciplinary ethical guidelines to assist health educators to serve the best interests of patients and clients are suggested.

Turning experience into academic learning. The APL and APEL credit systems.

1. Learning from experience can be turned into academic credit through APL and APEL. 2. The APL/APEL process is about valuing and empowering practitioners. 3. Reflection is central to developing an APEL claim. 4. An accreditation model offers a five-stage process leading to the award of credit.

Evidence-based practice: a retrograde step? The importance of pluralism in evidence generation for the practice of health care.

The origin of evidence-based medicine is explored and its connection to evidence-based practice examined. Widespread acceptance of the dominant scientific paradigm supporting evidence-based practice is challenged. Knowledge classifications and their relevance for nursing practice are considered, with the need for a critical balance to be pursued. Recommendations are made, regarding the future of evidence for practice, and a brief example of emerging evidence concerning values of professional caring is offered in redressing the balance.

Variation in the susceptibility of BHK populations and cloned cell lines to three strains of foot-and-mouth disease virus.

BHK monolayer and suspension cell populations maintained in different laboratories were found to vary in their susceptibility to infection with three strains of foot-and-mouth disease virus (FMDV). The susceptibility of parent cell populations was compared with that of individual clones of cells derived from them. The populations tested in this way consisted of a number of cell types, each expressing a different capacity to produce FMDV. The relative numbers of susceptible and insusceptible cells in each population appeared to determine the overall susceptibility of that population. The ability of the FMDV strain Asia 1 Iran 1/73 to multiply was an indicator of the general FMDV susceptibility of a BHK cell population.

Academic validation of prior and experiential learning: evaluation of the process.

This report is a summary of an AP(E)L evaluative research project conducted at the University of Southampton School of Nursing and Midwifery. The project aimed to shed light on the perceptions of candidates and teachers in conjunction with the effectiveness of the accreditation process. Through initial semi-structured interviews with teachers (n = 6) and candidates (n = 6) offering their perceptions and experiences, an emergent questionnaire was sent to the remaining active accreditation candidates (n = 60, response n = 39) and advisers/assessors (n = 36), response n = 32). The effectiveness of candidate preparation for beginning an accreditation claim, the comprehensiveness of the types of credit available to capture learning through experience, the styles of advising and feedback from assessment are all reported on. Interestingly, the data highlight how preparing an accreditation claim can affect both adviser and candidate, in terms of being more self-critical and reflective in their areas of practice.

A view of the phenomenon of caring in nursing practice.

Care is regularly used as a suffix to nursing, in such well-known phrases as 'total nursing care' and 'holistic nursing care'. While most care is provided by lay persons, there is little nursing research which focuses on the meaning of care, particularly in relation to the United Kingdom. This small-scale study investigates the meaning of care from the experience of six practising staff nurses in a British hospital and leads to a view of this phenomenon. Phenomenology was the chosen methodology, which facilitated the emergence of an essential structure of caring which incorporated four major categories described as 'being supportive', 'communicating', 'pressure' and 'caring ability'. It is suggested that, through gaining perspectives to enhance our understanding of the meaning of care, it will ultimately develop our understanding of nursing itself.

Mechanism of the 2,3-diphosphoglycerate-dependent phosphoglycerate mutase from rabbit muscle.

1. The properties and kinetics of the 2,3-diphosphoglycerate-dependent phosphoglycerate mutases are discussed. There are at least three possible mechanisms for the reaction: (i) a phosphoenzyme (Ping Pong) mechanism; (ii) an intermolecular transfer of phosphate from 2,3-diphosphoglycerate to the substrates (sequential mechanism); (iii) an intramolecular transfer of phosphate. It is concluded that these mechanisms cannot be distinguished by conventional kinetic measurements. 2. The fluxes for the different mechanisms are calculated and it is shown that it should be possible to distinguish between the mechanisms by appropriate induced-transport tests and by comparing the fluxes of (32)P- and (14)C-labelled substrates at chemical equilibrium. 3. With (14)C-labelled substrates no induced transport was found over a wide concentration range, and with (32)P-labelled substrates co-transport occurred that was independent of concentration over a twofold range. (14)C-labelled substrates exchange at twice the rate of (32)P-labelled substrates at chemical equilibrium. The results were completely in accord with a phosphoenzyme mechanism and indicated a rate constant for the isomerization of the phosphoenzyme of not less than 4x10(6)s(-1). The intramolecular transfer of phosphate (and intermolecular transfer between two or more molecules of substrate) were completely excluded. The intermolecular transfer of phosphate from 2,3-diphosphoglycerate would have been compatible with the results only if the K(m) for 2-phosphoglycerate had been over 7.5-fold smaller than the observed value and if an isomerization of the enzyme-2,3-diphosphoglycerate complex had been the major rate-limiting step in the reaction. 4. The very rapid isomerization of the phosphoenzyme that the experiments demonstrate suggests a mechanism that does not involve a formal isomerization. According to this new scheme the enzyme is closely related mechanistically and perhaps evolutionarily to a 2,3-diphosphoglycerate diphosphatase.

The mechanism of the phosphoglucomutase reaction. Studies on rabbit muscle phosphoglucomutase with flux techniques.

1. The kinetics of phosphoglucomutases from different sources are discussed and it is concluded that on the available evidence there are in all cases three possible mechanisms for the reaction. These are an indirect transfer of phosphate involving the phosphoenzyme (mechanism 1), a direct transfer of phosphate (mechanism 2), and an intermolecular transfer of phosphate from glucose 1,6-diphosphate to the substrate (mechanism 3). Conventional net flux measurements are shown not to differentiate between these mechanisms. 2. Flux equations are developed and it is shown that there are three flux ratios that characterize and distinguish between the mechanisms. 3. To examine these flux ratios induced-transport tests are described with (14)C- and (32)P-labelled substrates. The fluxes determined with (14)C- and (32)P-labelled substrates are also compared at chemical equilibrium. 4. With rabbit muscle phosphoglucomutase the results of these tests were completely consistent with mechanism 1 and unequivocally excluded any substantial part of the reaction proceeding by mechanism 2 or mechanism 3. Evidence was also obtained for an isomerization of the phosphoenzyme with an apparent rate constant about 4.5x10(7)sec.(-1). Taking into account the activity coefficients of the substrates the true rate constant appears to be about one-sixth of this value. 5. Isotope effects and non-ideal behaviour of the solutions are discussed and the activity coefficients of the substrates are shown to be equal by measurement of the depression of freezing point. It is concluded that these factors do not influence the tests significantly. 6. Alternative mechanisms are considered and it is concluded that the tests show that the glucose residue is transferred directly, that the phosphate is transferred indirectly with one intermediate phosphate, and that there is an isomerization of the free phosphoenzyme without reference to any other details of the reaction. Further, no assumptions are required about the constancy of rate constants. 7. The relative merits of induced transport and product inhibition for detecting isomerization of the enzyme are discussed. It is concluded that the induced-transport test is more sensitive and that its interpretation is less equivocal. 8. The application of the tests to other enzyme systems is briefly considered.

Mechanism of action of rabbit liver phosphoglucomutase.

Induced-transport tests with comparatively undegraded rabbit liver phosphoglucomutase show that the enzyme possesses a phosphoenzyme mechanism and that any interconversion of phosphoenzyme forms is very rapid. A relatively stable 32P-labelled phosphoenzyme was isolated, which exchanged label rapidly with substrates. The phospho group appears to be bonded to a serine residue on the enzyme.

Studies on the susceptibility to foot--and--mouth disease virus of BHK cell cultures derived from various sources.

Cultures of BHK monolayer and suspension cells obtained from a number of laboratories and a group of cloned sub-lines derived from suspension cells were examined for their susceptibility to three FMD virus strains. It was found that the various cultures were sensitive to the test virus strains to differing degrees. It was shown possible to obtain a clone with high susceptibility to Asia 1 Iran 1/73 virus from a parent culture which had a low susceptibility to that virus. The implication of these findings is discussed.