[Anesthesiological aspects of liver transplantation]. / Anästhesiologische Aspekte bei der Lebertransplantation.

Liver transplantation (LTPL) is the only curative option for patients with end stage liver disease (ESLD) or with hepatocellular carcinoma (HCC). Eurotransplant in Leiden, the Netherlands, is responsible for organ allocation. The model of end stage liver disease (MELD) score, which describes the severity of the liver disease, is decisive for organ allocation. The heterogeneous patient collective and hepatic-related comorbidities and their dynamics represent challenges. The anesthesiologist is responsible for evaluating the overall prognosis, whereby cardiac, pulmonary, renal and neurological comorbidities must be taken into consideration. During LTPL surgery is divided into several stages. Besides volume management, heat preservation and coagulation management, major challenges for the anesthesiologist are hemodynamic stabilization and regulation of the acid-base balance.

A comparison of single particle tracking and temporal image correlation spectroscopy for quantitative analysis of endosome motility.

Single particle tracking (SPT) is becoming a standard method to extract transport parameters from time-lapse image sequences of fluorescent vesicles in living cells. Another method to obtain these data is temporal image correlation spectroscopy (TICS), but this method is less often used for measurement of intracellular vesicle transport. Here, we present an extensive comparison of SPT and TICS. First we examine the effect of photobleaching, shading and noise on SPT and TICS analysis using simulated image sequences. To this end, we developed a simple photophysical model, which relates spatially varying illumination intensity to the bleaching propensity and fluorescence intensity of the moving particles. We found that neither SPT nor TICS are affected by photobleaching per se, but the transport parameters obtained by both methods are sensitive to the signal-to-noise ratio. In addition, the number of obtained trajectories in SPT is affected by noise. Diffusion constants determined by TICS are significantly overestimated when large immobile fluorescent structures are present in the image sequences, while the opposite is true for SPT. To improve the performance of both techniques, we compare three different methods for image denoising. Appropriate denoising significantly reduced the effect of noise and of immobile structures on both methods. Shape fluctuations of simulated particles had a more pronounced effect on TICS than on SPT analysis. In denoised images of fluorescent beads or cytosolic vesicles containing fluorescent protein NPC2 in human skin fibroblast cells, the transport parameters acquired by SPT and TICS were comparable emphasizing the value of both analysis methods.

Cellular effects and metabolic stability of N1-cyclic inosine diphosphoribose and its derivatives.

BACKGROUND AND PURPOSE: Recently, a number of mimics of the second messenger cyclic ADP-ribose (cADPR) with replacement of adenosine by inosine were introduced. In addition, various alterations in the molecule ranging from substitutions at C8 of the base up to full replacement of the ribose moieties still retained biological activity. However, nothing is known about the metabolic stability and cellular effects of these novel analogues. EXPERIMENTAL APPROACH: cADPR and the inosine-based analogues were incubated with CD38, ADP-ribosyl cyclase and NAD-glycohydrolase and metabolism was analysed by RP-HPLC. Furthermore, the effect of the analogues on cytokine expression and proliferation was investigated in primary T-lymphocytes and T-lymphoma cells. KEY RESULTS: Incubation of cADPR with CD38 resulted in degradation to adenosine diphosphoribose. ADP-ribosyl cyclase weakly catabolised cADPR whereas NAD-glycohydrolase showed no such activity. In contrast, N1-cyclic inosine 5'-diphosphoribose (N1-cIDPR) was not hydrolyzed by CD38. Three additional N1-cIDPR analogues showed a similar stability. Proliferation of Jurkat T-lymphoma cells was inhibited by N1-cIDPR, N1-[(phosphoryl-O-ethoxy)-methyl]-N9-[(phosphoryl-O-ethoxy)-methyl]-hypoxanthine-cyclic pyrophosphate (N1-cIDP-DE) and N1-ethoxymethyl-cIDPR (N1-cIDPRE). In contrast, in primary T cells neither proliferation nor cytokine expression was affected by these compounds. CONCLUSIONS AND IMPLICATIONS: The metabolic stability of N1-cIDPR and its analogues provides an advantage for the development of novel pharmaceutical compounds interfering with cADPR mediated Ca2+ signalling pathways. The differential effects of N1-cIDPR and N1-cIDPRE on proliferation and cytokine expression in primary T cells versus T-lymphoma cells may constitute a starting point for novel anti-tumor drugs.

Ochratoxin A producing Penicillium verrucosum isolates from cereals reveal large AFLP fingerprinting variability.

AIMS: To examine if molecular amplified fragment length polymorphism (AFLP) fingerprinting of the only ochratoxin A-producing species in European cereals, Penicillium verrucosum, can be used as a method in hazard analysis using critical control points (HACCP). METHODS AND RESULTS: A total of 321 isolates of P. verrucosum were isolated from ochratoxin A-contaminated cereals from Denmark (oats), UK (wheat and barley) and Sweden (wheat). Of these, 236 produced ochratoxin A as determined by thin layer chromatography; 185 ochratoxin A-producing isolates were selected for AFLP fingerprinting. A total of 138 isolates had unique AFLP patterns, whereas 52 isolates could be allocated to small groups containing from two to four isolates with similar AFLP patterns. A total of 155 clones were found among the 185 P. verrucosum isolates, thus 84% of the isolates may represent different genets of P. verrucosum. As the few isolates that were grouped often came from the same farm, and those groups that contained AFLP-identical isolates from different countries were morphotypically different. On single farms up to 35 clones were found. The few groups of ramets from the same genet indicated that a HACCP approach based on clones may require a very large number of AFLP analysis to work in practice, we recommend basing the HACCP approach on the actual species P. verrucosum. A more detailed characterization should rather be based on the profile of species present at different control points, or analysis of the mycotoxins ochratoxin A and citrinin in the isolates. Examination of 86 isolates with HPLC and diode array detection of P. verrucosum showed that 66% produced ochratoxin A, 87% produced citrinin, 92% produced verrucin and 100% produced verrucolone. CONCLUSIONS: Among 184 ochratoxin A-producing Penicillium verrucosum, 155 clonal lineages were indicated by AFLP fingerprinting, indicating a high genetical diversity, yet the species P. verrucosum is phenotypically distinct and valid. SIGNIFICANCE AND IMPACT OF THE STUDY: AFLP fingerprinting of Penicillium verrucosum indicates that genetic recombination takes place in this fungus.

Penicillium verrucosum in wheat and barley indicates presence of ochratoxin A.

AIMS: The aims of this study were to isolate and identify ochratoxin A (OTA) producing fungi in cereals containing OTA and to determine the best selective and indicative medium for recovery of OTA producing fungi. METHODS AND RESULTS: Seventy-six wheat, barley and rye samples from Europe containing OTA and 17 samples without OTA were investigated using three different media, dichloran yeast sucrose agar (DYSG), dichloran rose bengal yeast extract sucrose agar (DRYES) and dichloran 18% glycerol agar (DG18). Hundred kernels were plated on each medium and the kind and number of fungal OTA producers were recorded as percentage of infestation. Penicillium verrucosum was the sole OTA producer found in cereals. The average percentage of infestation of P. verrucosum counts was recorded as 28.3% on DYSG, 10.3% on DRYES and 9.9% on DG18 on the OTA containing samples and 0.8% on DYSG, 0.4% on DRYES and 0.6% on DG18 for the samples without OTA. CONCLUSIONS: Penicillium verrucosum was the sole OTA producer in European cereals. Determination of P. verrucosum infestation and infection was best detected on DYSG after 7 days at 20 degrees C. The percentage of infestation of P. verrucosum found on DYSG and OTA content in cereals were correlated. More than 7% infestation of P. verrucosum indicated OTA contamination. SIGNIFICANCE AND IMPACT OF THE STUDY: The developed method could be used as a cereal quality control.

Normal-mode-vortex interactions.

Standing surface waves that interact with a confined, vertical, vorticity field with zero net circulation are studied both analytically and experimentally. The surface waves are generated by vertical vibration, and constant vorticity injection is achieved by a rotating disk flush mounted in the cell. Experimental results are indicative of a local wave-vortex interaction (no dislocation), and a simple theoretical model is able to explain them in quantitative detail.

Cyclic ADP-ribose production by CD38 regulates intracellular calcium release, extracellular calcium influx and chemotaxis in neutrophils and is required for bacterial clearance in vivo.

Cyclic ADP-ribose is believed to be an important calcium-mobilizing second messenger in invertebrate, mammalian and plant cells. CD38, the best-characterized mammalian ADP-ribosyl cyclase, is postulated to be an important source of cyclic ADP-ribose in vivo. Using CD38-deficient mice, we demonstrate that the loss of CD38 renders mice susceptible to bacterial infections due to an inability of CD38-deficient neutrophils to directionally migrate to the site of infection. Furthermore, we show that cyclic ADP-ribose can directly induce intracellular Ca++ release in neutrophils and is required for sustained extracellular Ca++ influx in neutrophils that have been stimulated by the bacterial chemoattractant, formyl-methionyl-leucyl-phenylalanine (fMLP). Finally, we demonstrate that neutrophil chemotaxis to fMLP is dependent on Ca++ mobilization mediated by cyclic ADP-ribose. Thus, CD38 controls neutrophil chemotaxis to bacterial chemoattractants through its production of cyclic ADP-ribose, and acts as a critical regulator of inflammation and innate immune responses.

Ultrasound scattering and the study of vortex correlations in disordered flows

In an idealized way, some turbulent flows can be pictured by assemblies of many vortices characterized by a set of particle distribution functions. Ultrasound provides a useful, nonintrusive, tool to study the spatial structure of vorticity in flows. This is analogous to the use of elastic neutron scattering to determine liquid structure. We express the dispersion relation, as well as the scattering cross section, of sound waves propagating in a "liquid" of identical vortices as a function of vortex pair correlation functions. In two dimensions, formal analogies with ionic liquids are pointed out.

Computerized analysis of video fluorescein imaging (VFI) of the skin.

Video fluorescein imaging (VFI) is a new technique to continuously follow the development of fluorescence in the skin, i.e. blood inflow and perfusion, after intravenous injection of sodium fluorescein. The method is supplementary to other microcirculatory techniques for evaluation of peripheral arterial occlusive disease, particularly in critical ischaemia. In the present article we describe a totally computerized digital imaging processing system for evaluation and present results from a comparison between the evaluations of the appearance and development of the fluorescence in the sole of the foot using the computerized and the previously used manual techniques. With the computerized system the images are stored and correlated with the start of the injection. Regions of interest are then marked and a mean value of fluorescence intensity is calculated for each image. Using this computerized system the time required for evaluation has been shortened to about 10 min. The results of the comparison between the manual and computerized evaluations of appearance times showed that a significant correlation existed in all examined parts of the feet between the two techniques. The methods gave approximately the same results in regions with fluorescence appearance times between 20 and 50 s. With longer appearance times than approximately 50 s a systematic difference between the two techniques seemed to exist. In this interval shorter appearance times were measured with the computerized technique than with the manual technique. However, the clinical information with regard to prognosis would be relatively unchanged when the new computerized assessment technique and a new cut-off level for the appearance time are used. Also, regarding the development of fluorescence after the appearance time, expressed by the slope, a significant correlation was found between the manual and the computerized evaluation.

[Tuberculosis of the hand]. / Håndtuberkulose.

Since 1985 there has been an increase in the incidence of skeletal tuberculosis in Denmark. This increase is attributed to a high incidence of tuberculosis among HIV-positive patients, elderly people and the increasing number of immigrants from areas with endemic tuberculosis. We present a case of hand tuberculosis in a young Somali male adult, where the tubercle bacilli find their way to the third metacarpal bone of the left hand, without a history of trauma or previous pulmonary tuberculosis. There was a delay in the treatment due to misdiagnosis of just three weeks. The abscess totally vanished two months after start of treatment.

Chemical characterisation of cheese associated fungi.

Recent work in our laboratory has demonstrated that the most common contaminating fungi on different types of cheese are;Penicillium commune, P. nalgiovense, P. solitum, P. discolor, P. roqueforti, P. crustosum, P. nordicum andAspergillus versicolor. On blue cheese a new speciesP. caseifulvum has been discovered as a surface contaminant. A large number of known and unknown metabolites have been described from the above mentioned cheese associated fungi from both synthetic media and real samples. Based on chemotaxonomy our laboratory has discovered thatP. roqueforti should be divided into three species:P. roqueforti (from cheese),P. carneum (from meat) andP. paneum (from bread). SimilarlyP. verrucosum should be divided intoP. verrucosum (from cereals) andP. nordicum (from cheese and meat products). Both species produce ochratoxins, however, only the former species produce citrinin.

Reciprocal regulation of polarized cytokine production by effector B and T cells.

Although B cells produce cytokines it is not known whether B cells can differentiate into effector subsets that secrete polarized arrays of cytokines. We have identified two populations of "effector" B cells (Be1 and Be2) that produce distinct patterns of cytokines depending on the cytokine environment in which the cells were stimulated during their primary encounter with antigen and T cells. These effector B cell subsets subsequently regulate the differentiation of naïve CD4+ T cells to TH1 and TH2 cells through production of polarizing cytokines such as interleukin 4 and interferon gamma. In addition, Be1 and Be2 cells could be identified in animals that were infected with pathogens that preferentially induce a Type 1 and Type 2 immune response. Together these results suggest that, in addition to their well defined role in antibody production, B cells may regulate immune responses to infectious pathogens through their production of cytokines.

Intravenous hydroxyethylrutosides combined with long-term oral anticoagulation in atherosclerotic nonreconstructable critical leg ischemia: a retrospective study.

OBJECTIVE: To evaluate in a group of seriously diseased patients with nonreconstructable chronic critical leg ischemia (CLI), treated by a combination of i.v. hydroxyethylrutosides (HR)* and oral anticoagulation (AC) by warfarin, the short-term effects on the cutaneous microvascular blood perfusion of the soles of feet and especially the long-term clinical outcome in terms of amputation and death. DESIGN: A retrospective comparison between two groups of patients, HR + AC and a comparable reference group, fulfilling the same inclusion and exclusion criteria corresponding to the definition of CLI according to the Second European Consensus Document (1991). Clinical follow-up in both groups was made after 1, 3, 6, 12, and 24 months. SETTING: Patients were examined at university departments of clinical physiology with special interest in peripheral vascular disease, in cooperation with colleagues at university departments of surgery, internal medicine and dermatology of Karolinska Hospital, Södersjukhuset and Huddinge Hospital. PATIENTS: A total of seventy patients with CLI according to the definition of the Second European Consensus Document, 1991, ie, besides severe rest pain or ischemic lesions also a toe blood pressure < 30 mg Hg. Group with HR + anticoagulation (AC): 42 patients (19 diabetics, 23 nondiabetics). Reference group: 28 patients (18 diabetics, 10 nondiabetics). For distribution of age and toe blood pressure at baseline, see Table I. INTERVENTIONS: Therapy group: besides ordinary standard therapy, daily HR infusions for a mean period of 3.6 weeks + oral anticoagulation continued to the end of the study at 24 months. A comparable reference group on the same basic therapy but without the combination HR + AC. PARAMETERS IN EVALUATION: Short-term parameters: clinical data, skin temperature, and fluorescein imaging. Long-term outcome: amputation or death. RESULTS: Short-term and long-term results with HR + AC indicated that patients with severe CLI and very poor prognosis benefited in terms of survival and limb salvage from initial therapy with HR infusion combined with long-term oral anticoagulation. Results of this combined treatment seem at least comparable with those with i.v. prostacyclin analogies.

CD38 signaling in B lymphocytes is controlled by its ectodomain but occurs independently of enzymatically generated ADP-ribose or cyclic ADP-ribose.

CD38 is a type II transmembrane glycoprotein that is expressed by many cell types including lymphocytes. Signaling through CD38 on B lymphocytes can mediate B cell activation, proliferation, and cytokine secretion. Additionally, coligation of CD38 and the B cell Ag receptor can greatly augment B cell Ag receptor responses. Interestingly, the extracellular domain of CD38 catalyzes the conversion of NAD+ into nicotinamide, ADP-ribose (ADPR), and cyclic ADPR (cADPR). cADPR can induce intracellular calcium release in an inositol trisphosphate-independent manner and has been hypothesized to regulate CD38-mediated signaling. We demonstrate that replacement of the cytoplasmic tail and the transmembrane domains of CD38 did not impair CD38 signaling, coreceptor activity, or enzyme activity. In contrast, independent point mutations in the extracellular domain of CD38 dramatically impaired signal transduction. However, no correlation could be found between CD38-mediated signaling and the capacity of CD38 to catalyze an enzyme reaction and produce cADPR, ADPR, and/or nicotinamide. Instead, we propose that CD38 signaling and coreceptor activity in vitro are regulated by conformational changes induced in the extracellular domain upon ligand/substrate binding, rather than on actual turnover or generation of products.

Generalized Griffith criterion for dynamic fracture and the stability of crack motion at high velocities.

We use Eshelby's energy momentum tensor of dynamic elasticity to compute the forces acting on a moving crack front in a three-dimensional elastic solid [Philos. Mag. 42, 1401 (1951)]. The crack front is allowed to be any curve in three dimensions, but its curvature is assumed small enough so that near the front the dynamics is locally governed by two-dimensional physics. In this case the component of the elastic force on the crack front that is tangent to the front vanishes. However, both the other components, parallel and perpendicular to the direction of motion, do not vanish. We propose that the dynamics of cracks that are allowed to deviate from straight line motion is governed by a vector equation that reflects a balance of elastic forces with dissipative forces at the crack tip, and a phenomenological model for those dissipative forces is advanced. Under certain assumptions for the parameters that characterize the model for the dissipative forces, we find a second order dynamic instability for the crack trajectory. This is signaled by the existence of a critical velocity V(c) such that for velocities VV(c) it is governed by K(II) not equal to 0. This result provides a qualitative explanation for some experimental results associated with dynamic fracture instabilities in thin brittle plates. When deviations from straight line motion are suppressed, the usual equation of straight line crack motion based on a Griffiths-like criterion is recovered.

Scattering of dislocated wave fronts by vertical vorticity and the Aharonov-Bohm effect. I. Shallow water.

When a surface wave interacts with a vertical vortex in shallow water the latter induces a dislocation in the incident wave fronts that is analogous to what happens in the Aharonov-Bohm effect for the scattering of electrons by a confined magnetic field. In addition to this global similarity between these two physical systems there is scattering. This paper reports a detailed calculation of this scattering, which is quantitatively different from the electronic case in that a surface wave penetrates the inside of a vortex while electrons do not penetrate a solenoid. This difference, together with an additional difference in the equations that govern both physical systems, lead to a quite different scattering in the case of surface waves, whose main characteristic is a strong asymmetry in the scattering cross section. The assumptions and approximations under which these effects happen are carefully considered, and their applicability to the case of the scattering of acoustic waves by vorticity is noted.

Scattering of dislocated wave fronts by vertical vorticity and the Aharonov-Bohm effect. II. Dispersive waves.

Previous results on the scattering of surface waves by vertical vorticity on shallow water are generalized to the case of dispersive water waves. Dispersion effects are treated perturbatively around the shallow water limit, to first order in the ratio of depth to wavelength. The dislocation of the incident wave front, analogous to the Aharonov-Bohm effect, is still observed. At short wavelengths the scattering is qualitatively similar to the nondispersive case. At moderate wavelengths, however, there are two markedly different scattering regimes according to whether the depth is smaller or larger than sqrt[3] times capillary length. In the latter case, dispersion and advection may compensate leading to a spiral interference pattern. The dislocation is characterized by a parameter that depends both on phase and group velocity. The validity range of the calculation is the same as in the shallow water case: wavelengths small compared to vortex radius, and low Mach number. The implications of these limitations are carefully considered.

Mice deficient for the ecto-nicotinamide adenine dinucleotide glycohydrolase CD38 exhibit altered humoral immune responses.

CD38 is a membrane-associated ecto-nicotinamide adenine dinucleotide (NAD+) glycohydrolase that is expressed on multiple hematopoietic cells. The extracellular domain of CD38 can mediate the catalysis of NAD+ to cyclic adenosine diphosphoribose (cADPR), a Ca2+-mobilizing second messenger, adenosine diphosphoribose (ADPR), and nicotinamide. In addition to its enzymatic properties, murine CD38 has been shown to act as a B-cell coreceptor capable of modulating signals through the B-cell antigen receptor. To investigate the in vivo physiological function(s) of this novel class of ectoenzyme we generated mice carrying a null mutation in the CD38 gene. CD38-/- mice showed a complete loss of tissue-associated NAD+ glycohydrolase activity, showing that the classical NAD+ glycohydrolases and CD38 are likely identical. Although murine CD38 is expressed on hematopoietic stem cells as well as on committed progenitors, we show that CD38 is not required for hematopoiesis or lymphopoiesis. However, CD38-/- mice did exhibit marked deficiencies in antibody responses to T-cell-dependent protein antigens and augmented antibody responses to at least one T-cell-independent type 2 polysaccharide antigen. These data suggest that CD38 may play an important role in vivo in regulating humoral immune responses.

Arrest of B lymphocyte terminal differentiation by CD40 signaling: mechanism for lack of antibody-secreting cells in germinal centers.

Despite extensive research, the role of CD40 signaling in B cell terminal differentiation remains controversial. Here we show that CD40 engagement arrests B cell differentiation prior to plasma cell formation. This arrest is manifested at a molecular level as a reduction in mRNA levels of secretory immunoglobulin gene products such as mu(s) and J chain as well as the loss of the transcriptional regulator BLIMP-1. Furthermore, the inhibition of B cell differentiation by CD40 engagement could not be overcome by either mitogens or cytokines, but could be reversed by antibodies that interfere with the CD40/gp39 interaction. These data suggest that secretory immunoglobulin is not produced by B cells that are actively engaged by gp39-expressing T cells.