Stress stimulates production of catecholamines in rat adipocytes.

The sympathoadrenal system is the main source of catecholamines (CAs) in adipose tissues and therefore plays the key role in the regulation of adipose tissue metabolism. We recently reported existence of an alternative CA-producing system directly in adipose tissue cells, and here we investigated effect of various stressors-physical (cold) and emotional stress (immobilization) on dynamics of this system. Acute or chronic cold exposure increased intracellular norepinephrine (NE) and epinephrine (EPI) concentration in isolated rat mesenteric adipocytes. Gene expression of CA biosynthetic enzymes did not change in adipocytes but was increased in stromal vascular fraction (SVF) after 28 day cold. Exposure of rats to a single IMO stress caused increases in NE and EPI levels, and also gene expression of CA biosynthetic enzymes in adipocytes. In SVF changes were similar but more pronounced. Animals adapted to a long-term cold exposure (28 days, 4°C) did not show those responses found after a single IMO stress either in adipocytes or SVF. Our data indicate that gene machinery accommodated in adipocytes, which is able to synthesize NE and EPI de novo, is significantly activated by stress. Cold-adapted animals keep their adaptation even after an exposure to a novel stressor. These findings suggest the functionality of CAs produced endogenously in adipocytes. Taken together, the newly discovered CA synthesizing system in adipocytes is activated in stress situations and might significantly contribute to regulation of lipolysis and other metabolic or thermogenetic processes.

A simple, small and low cost permanent magnet design to produce homogeneous magnetic fields.

A new portable, pocket-size NMR probe based on a novel permanent magnet arrangement is presented. It is based on a Halbach-type magnet design which mimics the field of a spherical dipole by using cylindrical bar and ring magnets. The magnet system is made up of only three individual magnets, and most field calculations and optimisations can be performed analytically. A prototype system has been built using a set of small, off the shelf commercially available permanent magnets. Proton linewidths of 50 ppm FWHM could be achieved at a field strength of 1T. Calculations show that with custom-sized permanent magnets, linewidths of less than 1 ppm can be achieved over sample volumes of up to 1 mm3, which would in theory enable chemical shift resolved proton spectroscopy on mass-limited samples. But even with the achieved linewidth of 50 ppm, this can be a useful portable sensor for small amounts of liquid samples with restricted molecular mobility, like gels, polymers or high viscosity liquids.

A mobile one-sided NMR sensor with a homogeneous magnetic field: the NMR-MOLE.

A new portable NMR sensor with a novel one-sided access magnet design, termed NMR-MOLE (MObile Lateral Explorer), has been characterised in terms of sensitivity and depth penetration. The magnet has been designed to be portable and create a volume with a relatively homogeneous magnetic field, 15,000 ppm over a region from 4 to 16 mm away from the probe, with maximum sensitivity at a depth of 10 mm. The proton NMR frequency is 3.3 MHz. We have demonstrated that with this approach a highly sensitive, portable, unilateral NMR sensor can be built. Such a design is especially suited for the characterisation of liquids in situations where unilateral or portable access is required.

Towards a medically approved technology for alginate-based microcapsules allowing long-term immunoisolated transplantation.

The concept of encapsulated-cell therapy is very appealing, but in practice a great deal of technology and know-how is needed for the production of long-term functional transplants. Alginate is one of the most promising biomaterials for immunoisolation of allogeneic and xenogeneic cells and tissues (such as Langerhans islets). Although great advances in alginate-based cell encapsulation have been reported, several improvements need to be made before routine clinical applications can be considered. Among these is the production of purified alginates with consistently high transplantation-grade quality. This depends to a great extent on the purity of the input algal source as well as on the development of alginate extraction and purification processes that can be validated. A key engineering challenge in designing immunoisolating alginate-based microcapsules is that of maintaining unimpeded exchange of nutrients, oxygen and therapeutic factors (released by the encapsulated cells), while simultaneously avoiding swelling and subsequent rupture of the microcapsules. This requires the development of efficient, validated and well-documented technology for cross-linking alginates with divalent cations. Clinical applications also require validated technology for long-term cryopreservation of encapsulated cells to maintaining a product inventory in order to meet end-user demands. As shown here these demands could be met by the development of novel, validated technologies for production of transplantation-grade alginate and microcapsule engineering and storage. The advances in alginate-based therapy are demonstrated by transplantation of encapsulated rat and human islet grafts that functioned properly for about 1 year in diabetic mice.

[Experimental testing of a new coil design for endoluminal MRI applied to the pig stomach]. / Experimentelle Erprobung eines neuen Spulendesigns zur endoluminalen MRT am Beispiel des Schweinemagens.

PURPOSE: Experimental feasibility study of a new MR-Coil concept for enhanced visualization of the gastric wall. MATERIAL AND METHODS: The newly developed single-loop receiver coil for endoluminal imaging (Fraunhofer Institute, St. Ingbert, Germany) was evaluated in 4 explanted pig stomachs in a 1.5T MR unit (Siemens Symphony, Erlangen, Germany) with T1 w and T2 w MR sequences in three planes. The new coil consists of a foldable and self-expanding single loop coil (receiver coil) of a shape memory metal (nitinol). It was covered with a biocompatible material (silicone) to prevent direct contact of the wire with stomach tissue. The coil assumes a circular configuration with a diameter of 8 cm because of its memory metal properties. The flexible characteristics of the material used allow the passage through the instrument channel (13 mm diameter) of a specially designed MR-compatible endoscope. The purpose of our study was to assess feasibility of the coil design as a first step in developing a new endoluminal MRI-concept. Additionally the number and signal intensity of visible gastric wall layers were evaluated and findings were correlated with histopathological results of a pig stomach. RESULTS: The new coil concept was a feasible system in all 4 cases and showed good image quality for analysis. On T1 w images, 3 layers were visible in all cases, and on T2 w images 4 different gastric wall layers were seen in 2 cases. Due to histopathological correlation, the different gastric wall layers were identified as follows: mucosa, submucosa and muscularis propria if three layers were depicted; in cases of 4 visible wall layers, serosa and subserosa could be detected additionally. For each gastric wall layer, a distinct signal intensity was found. CONCLUSION: The new MR coil concept for endoluminal imaging proved to be a feasible technique. Good differentiation of gastric wall layers in the pig stomach could be demonstrated. We have shown that endoscopic MR-imaging with our new coil concept is a valuable technique for the visualization of gastric wall layers. Due to this fact, follow-up studies including assessing safety aspects are necessary to finally conduct an experimental-clinical study on in-vivo human gastric specimens to detect tumor growth and morphology within the gastric wall. Endoscopic MRI may have the potential in the future to overcome today's limitations of diagnostic imaging in gastric cancer.

Cross-linking properties of alginate gels determined by using advanced NMR imaging and Cu(2+) as contrast agent.

The entrapment of enzymes, drugs, cells or tissue fragments in alginates cross-linked with Ca(2+) or Ba(2+) has great potential in basic research, biotechnology and medicine. The swelling properties and, in turn, the mechanical stability are key factors in designing an optimally cross-linked hydrogel matrix. These parameters depend critically on the cross-linking process and seemingly minor modifications in manufacture have a large impact. Thus, sensitive and non-invasive tools are required to determine the spatial homogeneity and efficacy of the cross-linking process. Here, we show for alginate microcapsules (between 400 microm and 600 microm in diameter) that advanced (1)H NMR imaging, along with paramagnetic Cu(2+) as contrast agent, can be used to validate the cross-linking process. Two- and three-dimensional images and maps of the spin-lattice relaxation time T(1) of Ba(2+) cross-linked microcapsules exposed to external Cu(2+) yielded qualitative as well as quantitative information about the accumulation of Cu(2+) within and removal from microcapsules upon washing with Cu(2+) free saline solution. The use of Cu(2+) (having a slightly higher affinity constant to alginate than Ba(2+)) for gelling gave a complementary insight into the spatial homogeneity of the cross-linking process together with information about the mechanical stability of the microcapsules. The potential of this technique was demonstrated for alginates extracted from two different algal sources and cross-linked either externally by the conventional air-jet dropping method or internally by the "crystal gun" method.

Fabrication of homogeneously cross-linked, functional alginate microcapsules validated by NMR-, CLSM- and AFM-imaging.

Cross-linked alginate microcapsules of sufficient mechanical strength can immunoisolate cells for the long-term treatment of hormone and other deficiency diseases in human beings. However, gelation of alginate by external Ba(2+) (or other divalent cations) produces non-homogeneous cross-linking of the polymeric mannuronic (M) and guluronic (G) acid chains. The stability of such microcapsules is rather limited. Here, we show that homogeneous cross-linking can be achieved by injecting BaCl(2) crystals into alginate droplets before they come into contact with external BaCl(2). The high effectiveness of this crystal gun method is demonstrated by confocal laser scanning microscopy and by advanced nuclear magnetic resonance imaging. Both techniques gave clear-cut evidence that homogeneous cross-linkage throughout the microcapsule is only obtained with simultaneous internal and external gelation. Atomic force microscopy showed a very smooth surface topography for microcapsules made by the crystal gun method, provided that excess Ba(2+) ions were removed immediately after gelation. In vitro experiments showed greatly suppressed swelling for crystal gun microcapsules. Even alginate extracted from Lessonia nigrescens (highly biocompatible) yielded microcapsules with long-term mechanical stability not hitherto possible. Encapsulation of rat islets, human monoclonal antibodies secreting hybridoma cells and murine mesenchymal stem cells transfected with cDNA encoding for bone morphogenetic protein (BMP-4) revealed that injection of BaCl(2) crystals has no adverse side effects on cell viability and function. However, the release of low-molecular weight factors (such as insulin) may be delayed when using alginate concentrations in the usual range.

The impact of lipid distribution, composition and mobility on xylem water refilling of the resurrection plant Myrothamnus flabellifolia.

• Lipids play a crucial role in the maintenance of the structural and functional integrity of the water-conducting elements and cells of the resurrection plant Myrothamnus flabellifolia during complete dehydration. • Lipid composition, mobility and distribution within the internodal and nodal xylem regions (including short shoots and leaves) were investigated in the presence and absence of water by using various nuclear magnetic resonance (NMR) spectroscopy and imaging techniques differing greatly in the level of spatial resolution and acquisition of lipid parameters. • Significant findings include: a discontinuity in the branch xylem between an inner zone where no water moves and an outer zone where the water moves; the blocking of water movement in the inner zone by lipids that are not dispersed by water, and the facilitation of water advance in the xylem elements and pits of the outer zone by water-dispersed lipids; the relative impermeability of leaf trace xylem to the rehydrating water and, hence, the relative hydraulic isolation of the leaves. • These results elucidated part of the strategy used by the resurrection plant to cope with extreme drought and to minimize transpirational water loss upon hydration.

[ANCA-positive vasculitis of the skin and kidneys associated with acne conglobata]. / cANCA-positive Vaskulitis der Haut und Nieren im Verlauf einer Acne vulgaris conglobata.

A 19 year old man with severe acne conglobata and ulcerated pyoderma gangraenosum-like skin lesions on the face was first treated with isotretinoin (Roaccutan((R))), then immunosuppressively with prednisolone, diaminodiphenylsulfone (Dapson-Fatol((R))) and mycophenolate mofetil (Cellcept((R))). Under a daily maintenance dose of immunosuppressive treatment with 2.5 mg prednisolone and 1 g mycophenolate mofetil, weakness, muscle and joint aches appeared. Due to proteinuria, haematuria and an elevated antineutrophil cytoplasmic antibody (cANCA) and the histological detection of a leukocytoclastic vasculitis, the diagnosis of cANCA positive vasculitis of the skin and kidneys was established. Therapy with cyclophosphamide (Endoxan((R))) along with prednisolone was effective. An exact classification of this disease process was not possible.

Modeling categorical variables by logistic regression.

OBJECTIVE: To demonstrate the use of logistic regression in health care research. METHOD: Forward and backward stepwise logistic regression algorithms were systematically applied to a real-world data set comprising 301 cancer patients and a set of explanatory variables. RESULTS: Four variables were identified as effective predictors of pain reporting by cancer patients during chemotherapy: fatigue, depression, severity of colds or viral infections, and insomnia. The 4-predictor model was validated by (a) significance tests of regression coefficients at p<0.05, (b) significant improvement of this model over competing models, and (c) goodness of fit indices. CONCLUSIONS: Logistic regression is useful for health-related research in which outcomes of interest are often categorical.

Pain assessment in the cognitively impaired and unimpaired elderly.

The purpose of this study was to determine the self-report pain rating scale(s) that can be used to quantify pain in elderly persons across cognitive functioning levels. Randomly selected elderly subjects (N = 100) completed the Short Portable Mental Status Questionnaire to categorize their level of cognitive impairment: intact (n = 36), mild (n = 9), moderate (n = 15), and severe (n = 40). Pain was measured with the Memorial Pain Assessment Card verbal subscale, FACES, COOP pain subscale, a numeric rating scale, and the Present Pain Intensity subscale of the McGill Pain Questionnaire. Receiver operator characteristic curves indicated that participants categorized with moderate to no cognitive impairment were able to complete 1 or more of the pain assessment tools. Of the severely impaired, 30% were able to complete 1 or more pain assessment tools. Intraclass correlations showed a high degree of consistency among all pairs of tools (intraclass correlation > 0.74). We conclude that most elderly, with normal to moderately impaired cognitive functioning, as well as some severely impaired elderly, are capable of using self-report tools to rate their pain.

Echo-planar imaging of porous media with spatial resolution below 100 &mgr;m

A modified version of the echo-planar imaging technique incorporating a Carr-Purcell train of 180 degrees rf pulses (PEPI) has been implemented on a standard spectrometer. It is demonstrated that artifacts in the image due to cumulative errors in the rf field can be reduced by replacing each 180 degrees pulse by a composite sequence of three rf pulses. Artifact-free 3D images at 94 &mgr;m voxel resolution are obtained within 15 min. This technique has been applied to study the drying process in an initially water-saturated model porous medium with characteristic T*2 of order 700 &mgr;s. Copyright 1999 Academic Press.

Taylor dispersion and molecular displacements in Poiseuille flow.

We have used pulsed gradient spin echo (PGSE) NMR to measure longitudinal displacements of octane molecules undergoing Poiseuille flow in a 150 microm diameter pipe, accessing time scales which approach the Taylor dispersion limit. We monitor the change in displacement distribution which occurs as molecules undergoing Brownian motion sample an increasing proportion of the ensemble of streamlines, observing the effects of wall collisions and the gradual transition of the propagator from Poiseuille to Taylor-Aris behavior. The further use of a double PGSE sequence allows the direct measurement of the stochastic part of the motion alone.

Characterisation of fluid flow through porous media using three-dimensional microimaging and pulsed gradient stimulated echo NMR.

Pulsed gradient stimulated echo (PGSTE) and microimaging nuclear magnetic resonance (NMR) are used to probe correlations between structure and flow in the void space of a model porous system formed from a packing of 1-mm diameter glass spheres. The pulsed gradient stimulated echo data determine the average propagator and permit the dispersion of the flow to be studied as a function of delay time. Microimaging yields structural information and, specifically, a reduced radial distribution function (rdf) for the structure of the void space. Transition to fully developed dispersive flow is shown to occur on a scale size for which no further correlations in the structure of the void space are observed.

Absence of chronic effect of exposure to short-wave radio broadcast signal on salivary melatonin concentrations in dairy cattle.

A pilot study was conducted to investigate the influence of electromagnetic fields in the short-wave range (3-30 MHz) radio transmitter signals on salivary melatonin concentration in dairy cattle. The hypothesis to be tested was whether EMF exposure would lower salivary melatonin concentrations, and whether removal of the EMF source would be followed by higher concentration levels. For this pilot study, a controlled intervention trial was designed. Two commercial dairy herds at two farms were compared, one located at a distance of 500 m (exposed), the other at a distance of 4,000 m (unexposed) from the transmitter. At each farm, five cows were monitored with respect to their salivary melatonin concentrations over a period of ten consecutive days. Saliva samples were collected at two-hour intervals during the dark phase of the night. As an additional intervention, the short-wave transmitter was switched off during three of the ten days (off phase). The samples were analyzed using a radioimmunoassay. The average nightly field strength readings were 21-fold greater on the exposed farm (1.59 mA/m) than on the control farm (0.076 mA/m). The mean values of the two initial nights did not show a statistically significant difference between exposed and unexposed cows. Therefore, a chronic melatonin reduction effect seemed unlikely. However, on the first night of re-exposure after the transmitter had been off for three days, the difference in salivary melatonin concentration between the two farms (3.89 pg/ml, CI: 2.04, 7.41) was statistically significant, indicating a two- to seven-fold increase of melatonin concentration. Thus, a delayed acute effect of EMF on melatonin concentration cannot completely be excluded. However, results should be interpreted with caution and further trials are required in order to confirm the results.

Corticotropin-releasing hormone inhibits melatonin secretion in healthy volunteers--a potential link to low-melatonin syndrome in depression?

Interactions between the hypothalamic-pituitary-adrenocortical (HPA) system and melatonin secretion have been demonstrated, but only the effects of melatonin on the activity of the HPA system have been studied in man. Alterations of melatonin secretion described as low-melatonin syndrome have been demonstrated in patients suffering from a major depressive episode, and an inhibitory factor on melatonin secretion has been postulated. We investigated whether corticotropin-releasing hormone (CRH), which is thought to be involved in HPA abnormalities in depressed patients, can also suppress melatonin secretion in healthy volunteers. Ten healthy male human volunteers in a double-blind study design received randomized hourly intravenous injections from 08.00 to 18.00 h that contained 10 micrograms human CRH, 1 microgram adrenocorticotropic hormone (ACTH), or placebo to simulate pulsatile hormone secretion. Plasma melatonin and cortisol responses during the treatment and nocturnal sleep electroencephalograms after the treatment were recorded. Administration of CRH reduced melatonin secretion significantly below values obtained after administration of placebo and ACTH. Cortisol secretion was significantly enhanced by ACTH in comparison to both placebo and CRH. Electroencephalographic sleep parameters revealed no treatment effects. Our findings suggest that CRH has an inhibitory effect on the pineal secretion of melatonin in normal man. A mechanism via a release of cortisol was not supported by our results. Secondary hormonal effects from changes in nocturnal sleep architecture were excluded. Further investigation of the action of CRH on melatonin secretion as well as the mutual feedback between the HPA system and the pineal gland may extend our knowledge of neuroendocrine alterations mediating the adaptive response to stress and the eventual involvement in the pathogenesis of depression.