Solution structure of an oligodeoxynucleotide containing the human N-ras codon 12 sequence refined from 1H NMR using molecular dynamics restrained by nuclear Overhauser effects

A general model is developed for segmenting magnetic resonance images using vector decomposition and probability techniques. Each voxel is assigned fractional volumes of q tissues from p differently weighted images (q < or = p + 1) in the presence of partial-volume mixing, random noise, and other tissues. Compared with the eigenimage method, fewer differently weighted images are needed for segmenting the q tissues, and the contrast-to-noise ratio in the calculated fractional volumes is improved. The model can produce composite tissue-type images similar to that of the probability methods, by comparing the fractional volumes assigned to different tissues on each voxel. A three-tissue (p = 2, q = 3) model is illustrated for segmenting three tissues from dual-echo images. It provides statistical analysis to the algebraic method. A three-compartment phantom is segmented for validation. Two clinical examples are presented.     

Application of wavelet compression to digitized radiographs

Capillary electrophoresis is applied to determine nucleotide pool levels in human Burkitt lymphoma cells. The analysis was performed on a 65 cm x 50 microns I.D. Ucon-coated column with on-column UV detector. The method requires only nanoliters of sample and a simple sample preparation procedure. Over 12 nucleotides were separated and quantitated with high resolution and reproducibility. The whole capillary electrophoretic separation time was only 35 min. These results demonstrate that capillary electrophoresis provides a useful and easy way to analyze nucleotide pools in cells.     

The management of bereavement on intensive care units

OBJECTIVE: To investigate the management of the bereaved on Intensive Care Units (ICU) throughout the United Kingdom, and to identify inadequacies that may exist either in the provision of staff training in dealing with bereavement or in the facilities or support available for the bereaved. DESIGN: Questionnaires were sent to the senior nurse and senior doctor in all general ICUs with more than four beds nationwide. The questions asked about nursing and medical practice around the time of a patient's death, as well as about staff attitudes towards, and training in, dealing with bereavement and the support they received for this role. RESULTS: We obtained a 68% (293/430) response rate. Most ICUs had facilities for relatives, but little for the specific needs of the bereaved. Only 6% of doctors and 21% of nurses had training in dealing with bereavement and grieving. A staff support group was available in 23% of ICUs, and 75% of the remainder thought it would be useful to have one. Lack of staff training and poor facilities for relatives were identified as the major concerns of ICU staff. CONCLUSION: Many doctors and nurses working in Intensive Care Units feel inadequately trained to deal confidently with the bereaved. A minority of ICUs have support mechanisms available for their staff, inspite of the perceived need for them. Furthermore, many ICU staff feel the facilities they are able to offer the bereaved are inadequate. We have identified the major inadequacies and the needs of ICU staff for improved training. Meeting these needs would play a significant role not only in reducing staff stress but also minimising the morbidity in surviving relatives.     

Somatodendritic internalization and perinuclear targeting of neurotensin in the mammalian brain

Polypeptide hormones and growth factors have long been known to internalize into peripheral target cells as a result of their interaction with cell surface receptors. Studies in culture have suggested that certain neuropeptides might undergo a similar type of translocation in neurons. To investigate this possibility in adult mammalian brain, we have examined by confocal laser microscopy the events that follow the binding of fluorescein-tagged derivatives of the tridecapeptide neurotensin to basal forebrain cholinergic cells. Our results demonstrate a selective time- and temperature-dependent internalization of fluo-neurotensin in these cells. This internalization is receptor mediated, proceeds from the entire somatodendritic membrane of the cells, and utilizes endosome-like organelles which are mobilized from dendrites to perikarya and from the periphery of the cell to its perinuclear region. Parallel studies carried out on Sf9 insect cells expressing the rat neurotensin receptor from a recombinant baculovirus indicated that the internalization process involves receptor-ligand complexes and not merely the fluorescent peptide itself. These data suggest that receptor internalization plays a role in neuropeptide signaling in the brain and that it can be harnessed for selective identification of neuropeptide target cells.     

Identification of a kinetically distinct activity of 11beta-hydroxysteroid dehydrogenase in rat Leydig cells

Leydig cells are susceptible to direct glucocorticoid-mediated inhibition of testosterone biosynthesis but can counteract the inhibition through 11beta-hydroxysteroid dehydrogenase (11beta-HSD), which oxidatively inactivates glucocorticoids. Of the two isoforms of 11beta-HSD that have been identified, type I is an NADP(H)-dependent oxidoreductase that is relatively insensitive to inhibition by end product and carbenoxolone (CBX). The type I form has been shown to be predominantly reductive in liver parenchymal cells and other tissues. In contrast, type II, which is postulated to confer specificity in mineralocorticoid receptor (MR)-mediated responses, acts as an NAD-dependent oxidase that is potently inhibited by both end product and CBX. The identity of the 11beta-HSD isoform in Leydig cells is uncertain, because the protein in this cell is recognized by an anti-type I 11beta-HSD antibody, but the activity is primarily oxidative, more closely resembling type II. The goal of the present study was to determine whether the kinetic properties of 11beta-HSD in Leydig cells are consistent with type I, type II, or neither. Leydig cells were purified from male Sprague-Dawley rats (250 g), and 11beta-HSD was evaluated in Leydig cells by measuring rates of oxidation and reduction, cofactor preference, and inhibition by end product and CBX. Leydig cells were assayed for type I and II 11beta-HSD and MR messenger RNAs (mRNAs), and for type I 11beta-HSD protein. Leydig cell 11beta-HSD had bidirectional catalytic activity that was NADP(H)-dependent. This is consistent with the hypothesis that type I 11beta-HSD is present in rat Leydig cells. However, unlike the type I 11beta-HSD in liver parenchymal cells, the Leydig cell 11beta-HSD was predominantly oxidative. Moreover, analysis of kinetics revealed two components, the first being low a Michaelis-Menten constant (Km) NADP-dependent oxidative activity with a Km of 41.5 +/- 9.3 nM and maximum velocity (Vmax) of 7.1 +/- 1.2 pmol x min x 10(6) cells. The second component consisted of high Km activities that were consistent with type I:NADP-dependent oxidative activity with Km of 5.87 +/- 0.46 microM and Vmax of 419 +/- 17 pmol x min x 10(6) cells, and NADPH-dependent reductive activity with Km of 0.892 +/- 0.051 microM and Vmax of 117 +/- 6 pmol x min x 10(6) cells. The results for end product and CBX inhibition were also inconsistent with a single kinetic activity in Leydig cells. Type I 11beta-HSD mRNA and protein were both present in Leydig cells, whereas type II mRNA was undetectable. We conclude that the low Km NADP-dependent oxidative activity of 11beta-HSD in Leydig cells does not confirm to the established characteristics of type I and may reside in a new form of this protein. We also demonstrated the presence of the mRNA for MR in Leydig cells, and the low Km component could allow for specificity in MR-mediated responses.     

Fusion of the nucleoporin gene NUP98 to HOXA9 by the chromosome translocation t(7;11)(p15;p15) in human myeloid leukaemia

The parasympathetic nervous system plays a major role in the pathophysiology of many cardiovascular disease, particularly in modulating myocardial electrical stability. Measurements of heart rate variability have been widely used to assess parasympathetic activity. The reproducibility of measurements obtained from 24-hour ambulatory electrocardiograms has not been well documented. We have developed a technique for measuring parasympathetic activity from clinical quality 24-hour ambulatory electrocardiograms by counting beat-to-beat increases in RR interval that are > 50 ms. To determine the reproducibility and sensitivity of our technique, we analyzed repeated 24-hour electrocardiograms of 173 subjects (19 normal subjects, 67 patients with ischemic heart disease, and 87 diabetics) followed up over periods of 2 to 16 weeks. In all subject groups, mean values for repeated measurements were virtually identical. Measurements were stable in all 3 groups throughout the course of the study, as assessed by intraclass correlation coefficients. This technique is sensitive enough to detect relatively small changes in parasympathetic activity in subjects, as demonstrated by the calculated Bland and Altman coefficients of repeatability. Reproducibility and sensitivity of our technique are particularly good in normal subjects and in patients with ischemic heart disease. The results obtained with this technique imply that other related measurements of parasympathetic activity will show similar excellent short- and long-term reproducibility and sensitivity.