Kinase dynamics. Using ancient protein kinases to unravel a modern cancer drug's mechanism.

Macromolecular function is rooted in energy landscapes, where sequence determines not a single structure but an ensemble of conformations. Hence, evolution modifies a protein's function by altering its energy landscape. Here, we recreate the evolutionary pathway between two modern human oncogenes, Src and Abl, by reconstructing their common ancestors. Our evolutionary reconstruction combined with x-ray structures of the common ancestor and pre-steady-state kinetics reveals a detailed atomistic mechanism for selectivity of the successful cancer drug Gleevec. Gleevec affinity is gained during the evolutionary trajectory toward Abl and lost toward Src, primarily by shifting an induced-fit equilibrium that is also disrupted in the clinical T315I resistance mutation. This work reveals the mechanism of Gleevec specificity while offering insights into how energy landscapes evolve.

A qualitative study exploring perceptions and experiences of patients and clinicians of palliative medicine outpatient clinics in different settings.

Palliative care exists in a variety of settings and palliative care teams form many guises within this. A Palliative Medicine Outpatient Clinic (PMOC) exists to meet the flexible provision of the needs and preferences of individuals within whatever care setting they reside. This explorative study used a qualitative methodology, capturing patients' actual experience of care in preference to their satisfaction, as this is a more accurate measure of how and what patients judge as important in their healthcare. The overall themes in this paper point to the 'value' that patients perceived from attending the PMOC and how important the clinics were to clinicians that provided the care. The clinic facilitates much more than symptom control and here lies the challenge in how we convert the very positive experience of individuals into a language of outcome measures that captures the 'essence' of our work in this fiscally driven health economy.

Swallowing problems and dementia in acute hospital settings: practical guidance for the management of dysphagia.

There is little evidence to support tube feeding in advanced dementia although it is still frequently used in acute hospital settings. Patients present with complex problems and are often unable to make decisions about their healthcare needs. Multidisciplinary teams may be challenged by the difficult ethical decisions they are required to make in the best interests of their patients. This paper guides decision making in the management of patients with dementia and dysphagia in the acute hospital setting. A structured approach to information gathering, assessment and management is outlined with practical application of recent published guidelines and research evidence, which ensures appropriate individualised care.

Application of high-throughput technologies to a structural proteomics-type analysis of Bacillus anthracis.

A collaborative project between two Structural Proteomics In Europe (SPINE) partner laboratories, York and Oxford, aimed at high-throughput (HTP) structure determination of proteins from Bacillus anthracis, the aetiological agent of anthrax and a biomedically important target, is described. Based upon a target-selection strategy combining ;low-hanging fruit' and more challenging targets, this work has contributed to the body of knowledge of B. anthracis, established and developed HTP cloning and expression technologies and tested HTP pipelines. Both centres developed ligation-independent cloning (LIC) and expression systems, employing custom LIC-PCR, Gateway and In-Fusion technologies, used in combination with parallel protein purification and robotic nanolitre crystallization screening. Overall, 42 structures have been solved by X-ray crystallography, plus two by NMR through collaboration between York and the SPINE partner in Utrecht. Three biologically important protein structures, BA4899, BA1655 and BA3998, involved in tRNA modification, sporulation control and carbohydrate metabolism, respectively, are highlighted. Target analysis by biophysical clustering based on pI and hydropathy has provided useful information for future target-selection strategies. The technological developments and lessons learned from this project are discussed. The success rate of protein expression and structure solution is at least in keeping with that achieved in structural genomics programs.

Precision in transient electric birefringence measurements for colloids.

Electrically induced birefringence is increasingly used as a fast procedure to characterise the size, shape, polarisation and charge parameters of colloids and their interactions. By considering the optical apparatus generally used, attention is drawn to the significant errors that can arise in such experiments if optical component selection and setting are not critically considered.

European experience with a novel noninvasive sensor for intra-amniotic or extra-amniotic evaluation of fetal oxygen saturation.

OBJECTIVE: We evaluated the feasibility of a new instrument for continuous fetal pulse oximetry during labor. The measuring sensor can be placed on the fetal back before or after rupture of membranes. METHODS: One hundred adult women who had completed 32 weeks of gestation and had an anticipated duration of labor greater than 30 minutes were included in the study. Patients with premature rupture of membranes for 24 hours or more, low placental localization, placenta previa or abruption, vaginal bleeding, acute infection, polyhydramnios, oligohydramnios, or uterine or congenital abnormalities were excluded. RESULTS: All sensors were placed successfully. The mean continuous recording time was 276 minutes. Peripheral oxygen saturation as measured by pulse oximeter values were obtained during a median of 64.05% of the recording time. No chorioamnionitis or endometritis was noted. CONCLUSION: The new sensor instrumentation was safe for mother and fetus and was well accepted by parents and physicians.

Application of micro-scale sealed vessel thermal desorption-gas chromatography-mass spectrometry for the organic analysis of airborne particulate matter: linearity, reproducibility and quantification.

Micro-scale sealed vessel thermal desorption-gas chromatography-mass spectrometry (MSSV-TD-GC-MS) has been applied to the analysis of airborne particulate matter using the US NIST Standard Reference Material (SRM1649a) urban dust. We make qualitative comparisons with open system desorption and illustrate that caution should be used when using the technique without an open system comparison. We report linear responses over the same particulate mass range (1-5 mg) and good reproducibility [SD< or =0.62 mg kg(-1) (< or = 11%)]. We show linearity for a series of 10 n-alkanes and 10 polycyclic aromatic hydrocarbons (PAHs) in the SRM. The technique is also shown to be quantifiable (PAH concentrations typically 4-6 mg kg(-1)).

Quantification of polycyclic aromatic hydrocarbons in the NIST standard reference material (SRM1649A) urban dust using thermal desorption GC/MS

A thermal desorption GC/MS technique has been developed for the quantification of polycyclic aromatic hydrocarbons (PAHs) in airborne particulate matter using the NIST Standard Reference Material (SRM1649a) Urban Dust. The technique was developed using standard linearity tests in order to establish optimum sample weights and optimum desorption and chromatographic parameters. This direct analysis technique eliminates the use of solvents in the sample preparation (reducing volatile component losses) and also significantly reduces the sample preparation time (no extraction procedure). The technique has been shown to give linearity in terms of the overall TIC response as well as for a prominent series of n-alkanes (C20-C33) and 10 NIST priority PAHs, 8 of which have been quantified. The technique is reported to be uniquely sensitive (PAH concentrations 2-6 mg kg(-1)) and reproducible (MW = 178-228 SD < or =0.228 mg kg(-1), < or =7%; MW = 252 SD < or =0.922 mg kg(-1), < or =33%) over the range of sample weights (1-5 mg). Such sample weights illustrate that the technique can be equally applied to the analysis of airborne particulate samples collected over short time periods (24-48 h) using only commonly used low-volume collection devices.

Effect of bovine somatotropin on reproductive function in lactating dairy cows.

Several recent experiments have reported that chronic treatment with bovine somatotropin (bST) increased the number of days open without affecting the services per conception. The physiological basis for these effects was examined. Eleven lactating Holstein cows received daily injections of bST (40 mg) and 10 received daily injections of vehicle. Treatment was initiated between 32 and 85 d post partum and continued for up to 180 d. Eight of 11 bST-treated cows experienced at least one period of extended ovarian acyclicity during treatment. Only 3 of 10 control cows did so (P = 0.05). Concentrations of progesterone during luteal phases were lower in bST-treated cows than in controls (P = 0.06). Baseline concentrations of LH were suppressed in bST-treated cows compared with those of controls (P < 0.04). Neither the pulse frequency of LH nor the expression of estrous behavior was affected by bST (P > 0.30). These results indicate that chronic administration of a high dose of bST can reduce reproduction performance by promoting ovarian acyclicity.

Copper status and requirement during the dry period and early lactation in multiparous Holstein cows.

Eighteen multiparous Holstein cows were assigned randomly to three treatments at the beginning of the dry period (8 wk before expected parturition). Treatments were: 1) the basal diet containing 5.5 ppm of Cu (control), 2) the basal diet supplemented with 10 ppm of Cu, and 3) the basal diet supplemented with 20 ppm of Cu. The objectives were to measure the changes of Cu and other trace mineral element concentrations in blood and liver from the onset of the dry period (approximately 8 wk prepartum) to 8 wk postpartum and to assess the requirement of Cu during this time. Liver Cu concentration in the control group declined continuously during the 8-wk dry period, and the nadir occurred at parturition. This decline was prevented by dietary Cu supplementation of 10 or 20 ppm. Liver Cu concentration in the control group started to increase slowly after the dramatic decline. Liver Zn concentration changed cubically as a function of week during the treatment period. Plasma Cu and Zn exhibited a quadratic pattern as a function of week. The plasma Cu concentration was lowest 5 wk prior to parturition.

Symposium: Dream research methodology: Methodological issues affecting the collection of dreams.

This article focuses attention on the perennial issue of home versus laboratory recording situations for the garnering of dream reports. Studies into the experimental control over procedures for both waking and interviewing are reviewed. Against this background, an experiment into the relationship between REM/NREM sleep and visual imagery is reported. We found that, having corrected for total word count, visual imagery still made an independent contribution to REM reports. This finding is evaluated in the light of differential cognitive mechanisms underlying dream formation during sleep.

EEG spectral power analysis of phasic and tonic REM sleep in young and older male subjects.

EEG spectral power was studied during periods of rapid eye movements (REMs) and tonic intervals in REM sleep of 7 young and 7 older male subjects. Significant symmetrical decreases in alpha and beta1 power at central and occipital sites, concurrent with an increase in frontal theta power, were observed during the production of REMs. The former findings are discussed as sleep analogues to changes in alpha and beta1 during waking, showing increased information processing and behavioural activation, and that of theta is tentatively presented as reflecting an increase in afferent thresholds. Independent of the phasic-tonic REM distinction, total EEG power markedly decreased as a function of time of night and did not interact with age. Significant age differences in the overall spectral composition of the EEG were obtained, namely, a lower level of delta power and a relative shift towards more power in frequencies above 12 Hz for the older group. Further, older subjects also demonstrated a more uniform topographical distribution of alpha and sigma power.

Effect of copper status on luteinizing hormone secretion in dairy steers.

Two experiments were conducted to examine the effects of Cu status on neuroendocrine regulation of LH secretion in dairy steers. In Experiment 1, 18 Holstein steers were assigned randomly to treatment groups in a completely randomized design. Treatments consisted of basal diet alone (control) or supplemented with 5 ppm of Mo or with 20 ppm of Cu. At 8 and 16 wk, Cu concentrations in liver were higher in the group receiving Cu and lower in the group receiving Mo than in the control. Pulse frequencies of LH were not affected by any treatment. Serum LH concentration at 16 wk tended to be lower in the group receiving Mo than in either the control or the group receiving Cu. The ability of the pituitary gland to release LH in response to GnRH was not affected at 17 wk. Concentrations of LH in the pituitary glands were lower in the group receiving Mo than in that receiving Cu or in the controls. No differences in Cu or Mo concentrations were observed in pituitary, median eminence, or hypothalamic tissues. In Experiment 2, 12 Holstein steers were assigned to treatment groups receiving the basal diet supplemented with either 20 ppm or Cu or with 10 ppm of Mo. Spontaneous and GnRH-induced secretion of LH were not influenced by treatments after 8 mo of experiment. Reduction in Cu status has little effect on the endogenous secretion of LH in dairy steers.

Removal of ocular artifacts from the REM sleep EEG.

The present report concerns the first study in which electrooculographic (EOG) contamination of electroencephalographic (EEG) recordings in rapid eye movement (REM) sleep is systematically investigated. Contamination of REM sleep EEG recordings in six subjects was evaluated in the frequency domain. REM-active and REM-quiet series were obtained for each subject. Transfer coefficients and power spectra of EOG and EEG indicated that (a) increases in transfer coefficients beyond 4.5 Hz are brought about by residual EEG in the EOG, and (b) EOG-EEG contamination in the delta band is most pronounced in frontal, intermediate in central and negligible in occipital leads. It was found that correction of the REM-active series resulted in significant (c) reductions in power, (d) increases in interhemispheric coherences and (e) reductions in degree of lateral asymmetry. These effects were largest for frontal leads, but still marked for central ones. The results are discussed in the light of previous findings concerning models of hemispheric functioning during REM sleep.

Aging and memory for dreams.

This study investigated the influence of aging on memory for home drams and the extent to which cognitive variables such as visual memory, visuospatial IQ, and verbal IQ could account for possible differences herein. Subjects were 80 men and women of ages 45 to 75 years. Memory for dreams was measured by narrative length and frequency of recall. With respect to both measures no significant age differences were noted. Over-all differences in dream recall seemed best explained by visual memory scores. Partial correlational analyses, however, indicated that the small age differences memory or on any of the other cognitive variables. Dream contents were scored for aggression, friendliness, emotion, activities, and the number of characters and objects. The incidence of emotions among women appeared to be lower beyond the age of 60. Comparisons with previous data for young adults indicated that large reductions in aggression, friendliness, and emotion occur before the age of 45.

Effects of copper status on neutrophil function, superoxide dismutase, and copper distribution in steers.

Twelve Holstein steers in a completely randomized block design were fed either a basal diet (concentrate:silage or hay at a DM ratio of 35:65) plus Cu sulfate at 20 ppm of Cu (Cu-supplemented diet) or a basal diet plus ammonium molybdate to obtain 10 ppm of Mo (Cu-depleting diet) on a DM basis in the whole diet for 8 mo. Supplemental Mo was utilized in the Cu-depleting diet to develop a Cu-deficient group. Molybdenum slowly accumulated in the liver in the group fed the Cu-depleting diet. Copper concentrations in the liver and polymorphonuclear neutrophils decreased in the Cu-deficient group compared with the Cu-sufficient group. Plasma Cu concentration did not change during the trial for the Cu-sufficient group. In the Cu-deficient group, plasma Cu concentrations increased during the first 3 mo of the trial, then declined, and remained unchanged for the last 5 mo. Superoxide dismutase activities in red blood cells, polymorphonuclear neutrophils, and whole blood decreased in the Cu-deficient group. Phagocytic capacity was not affected by Cu status, but killing capacity was decreased by low Cu status in the Cu-deficient group by the end of the trial. Glutathione peroxidase activity was unaffected by Cu status. Clinical symptoms of Cu-deficiency were not observed in this trial; there was no evidence of blood hemoglobin or BW gain difference between the two groups. In this study, Cu status affected its distribution in the tissues and related enzyme activities as well as bactericidal function of neutrophils.

Effects of copper sources and dietary cation-anion balance on copper availability and acid-base status in dairy calves.

Twenty-four Holstein and Jersey calves (14 Holstein), 4 to 11 d of age, were assigned randomly to six treatments in a 2 x 3 factorial arrangement to examine the effects of Cu sources and dietary cation-anion balance on Cu availability and acid-base balance. Treatments were cationic basal diet (20 meq of dietary cation-anion balance on a DM basis), cationic basal diet supplemented with CuO, cationic basal diet supplemented with CuSO4, anionic basal diet (-10 meq), anionic basal diet supplemented with CuO, and anionic basal diet supplemented with CuSO4. Copper sources did not show any effect on growth of calves. The cationic diet increased calf growth compared with the anionic diet at wk 12 of the experiment. Blood pH was increased by the cationic diet in comparison with the anionic diet at wk 8 and 12. Blood pH also was increased by CuSO4 compared with CuO treatment in the early period of the treatment. Blood bicarbonate concentration was decreased by CuO and the anionic diet. Interactions between Cu sources and cation-anion balance were found for blood pH and bicarbonate concentration. Liver Cu concentration was increased by CuSO4 but not by CuO supplementation compared with control. Therefore, CuSO4 was found to be highly available, whereas CuO was a very poorly available source of Cu for young calves.

Role of magnesium in the dietary cation-anion balance equation for ruminants.

Eight midlactation Holstein cows (four primiparous) were assigned to replicated 4 x 4 Latin squares with a 2 x 2 factorial arrangement of treatments. The basal diet was formulated to contain 0 milliequivalents (Na + K) - Cl/100 g of diet DM. Treatment diets of high and low dietary cation-anion balance were achieved by adding appropriate amounts of supplemental Na + K (added on an equivalent basis) or Mg to the basal diet. Milk and protein yield were increased as well as DMI and FCM for Na + K diets The high concentration of cation increased milk fat percentage. The cation-anion balance (using Mg as a cation) had no effect on any of the production parameters observed. The higher cation-anion balances increased blood bicarbonate levels and plasma Na. Urinary Ca excretion was increased for the Mg diets. Urinary pH increased with both the source of cation (Na + K) and the concentration of cation (high) in the diet. other minerals were unaffected in milk, plasma, or urine. Responses measured in this trial reflect treatments calculated using the equation milliequivalents (Na + K) - Cl/100 g diet DM or (Na + K) - (Cl + S)/100 g diet DM, suggesting that Mg plays a minor role in dietary cation-anion balance.

Role of sulfur and chloride in the dietary cation-anion balance equation for lactating dairy cattle.

Ten Holstein cows averaging 120 d in lactation were arranged in replicated 5 x 5 Latin squares with 3-wk periods to evaluate the role of sulfur (S) in the dietary cation-anion balance equation. Diets were based on corn silage in Exp. 1 and sorghum silage in Exp. 2. Supplemental S and chloride (Cl) from the double sulfate of potassium and magnesium and CaCl2 were used to manipulate dietary cation-anion balance from 0 to +30 meq when expressed as meq [(Na + K)-(Cl + S)]/100 g diet DM and from +19 to +49 meq when expressed as meq [(Na + K)-Cl]/100 g diet DM. Blood pH was not affected by cation-anion balance, although both S and Cl supplementation tended to lower pH. Blood HCO3- and urine pH decreased and plasma calcium (Ca) and urinary Ca excretion increased as anion was added to the diet. Milk fat production tended to be increased by the low S supplementation. Dietary Cl and S had similar effects on acid-base status. Therefore, we suggest that S be included with Cl in the dietary cation-anion balance equation for lactating dairy cows as follows: meq [(Na + K)-(Cl + S)]/100 g diet DM. Although response of acid-base status to S and Cl was similar, as more data comparing the acidogenicity of S vs Cl become available, it may be necessary to include a modifying coefficient for S in the equation to adjust for differences between S and Cl in acid-generating potential. This coefficient may be further dependent on the dietary source of S.

Two siblings with Tel Hashomer camptodactyly and mitral valve prolapse.

A brother and sister with Tel Hashomer camptodactyly and mitral valve prolapse are described. Mitral valve prolapse is heterogenous, but appears to occur more frequently in individuals with connective tissue disorders. The presence of mitral valve prolapse as a component manifestation of Tel Hashomer camptodactyly suggests that abnormal connective tissue is a pleiotropic effect of the mutant allele.