Conformational dynamics of the tetracycline-binding aptamer

The conformational dynamics induced by ligand binding to the tetracycline-binding aptamer is monitored via stopped-flow fluorescence spectroscopy and time-correlated single photon counting experiments. The fluorescence of the ligand is sensitive to changes within the tertiary structure of the aptamer during and after the binding process. In addition to the wild-type aptamer, the mutants A9G, A13U and A50U are examined, where bases important for regulation are changed to inhibit the aptamer’s function. Our results suggest a very fast two-step-mechanism for the binding of the ligand to the aptamer that can be interpreted as a binding step followed by a reorganization of the aptamer to accommodate the ligand. Binding to the two direct contact points A13 and A50 was found to occur in the first binding step. The exchange of the structurally important base A9 for guanine induces an enormous deceleration of the overall binding process, which is mainly rooted in an enhancement of the back reaction of the first binding step by several orders of magnitude. This indicates a significant loss of tertiary structure of the aptamer in the absence of the base A9, and underlines the importance of pre-organization on the overall binding process of the tetracycline-binding aptamer

The neuropathology of fatal encephalomyelitis in human Borna virus infection

After many years of controversy, there is now recent and solid evidence that classical Borna disease virus 1 (BoDV-1) can infect humans. On the basis of six brain autopsies, we provide the first systematic overview on BoDV-1 tissue distribution and the lesion pattern in fatal BoDV-1-induced encephalitis. All brains revealed a non-purulent, lymphocytic sclerosing panencephalomyelitis with detection of BoDV-1-typical eosinophilic, spherical intranuclear Joest-Degen inclusion bodies. While the composition of histopathological changes was constant, the inflammatory distribution pattern varied interindividually, affecting predominantly the basal nuclei in two patients, hippocampus in one patient, whereas two patients showed a more diffuse distribution. By immunohistochemistry and RNA in situ hybridization, BoDV-1 was detected in all examined brain tissue samples. Furthermore, infection of the peripheral nervous system was observed. This study aims at raising awareness to human bornavirus encephalitis as differential diagnosis in lymphocytic sclerosing panencephalomyelitis. A higher attention to human BoDV-1 infection by health professionals may likely increase the detection of more cases and foster a clearer picture of the disease

Wirkung von Renin auf Kardiomyozyten der Ratte unter prohypertrophen Bedingungen

Einleitung: Die moykardiale Hypertrophie spielt eine wichtige Rolle in der Pathophysiolgie der Herzinsuffizienz. Dieser (mal)adaptive Mechanismus führt letztlich zum nekrotischen und apoptotischen Untergang von Herzmuskelgewebe, sowie zum bindegewebigen Umbau des Myokards mit Einschränkung der myokardialen Pumpleistung. Einen wichtigen Beitrag zur myokardialen Hypertrophie leistet das Renin-Angiotensin-(Aldosteron)-System (RA(A)S), welches durch seine Effektorhormone Angiotensin II und Aldosteron zum einen zur Erhöhung des Blutdrucks und zum anderen zu myokardialen Umbauprozessen beiträgt. Dies bietet auch zahlreiche Angriffspunkte für die aktuelle medikamentöse Herzinsuffizienztherapie. Für Renin, das Schlüsselenzym des RA(A)S, wurden mit dem spezifischen (Pro-)Renin-Rezeptor (PRR) und dem Mannose-6-Phosphat/Insulin-like growth factor II Rezeptor (M6P/IGFIIR) in den letzten Jahren mehrere Rezeptoren entdeckt. Über diese kann das Renin in die Zelle aufgenommen werden und auch hormonelle Wirkungen vermitteln. Auch die Renin-Vorstufe Prorenin kann an diese Rezeptoren binden, wird internalisiert und zum enzymatisch aktiven Renin umgewandelt. Die vorliegende Studie untersucht Wirkungen von Renin auf Kardiomyozyten der Ratte, welche unter prohypertrophem Stress stehen. Methoden: Kardiomyozyten männlicher Wistar-Ratten wurden in Kurzzeitkultur genommen und mit Renin in verschiedenen Konzentrationen sowie den bekannt prohypertrophen Stimulanzien FCS und Endothelin-1 inkubiert. Nach Inkubation über Nacht wurden die Zellen unter dem Mikroskop in Länge und Breite vermessen. Zudem wurden Parameter der lastfreien Zellverkürung unter der Zeilenkamera erhoben. Zur Übertragung der Ergebnisse auf ein Rattenmodell ex vivo wurden die Kardiomyozyten von Sprague-Dawley-Ratten, spontan hypertensiven Ratten und Renin-überexprimierenden transgenen Ratten des Typs TGR(mRen2)27 in Kurzzeitkultur genommen und den gleichen Untersuchungen unterzogen. Vor der Tötung der Tiere wurden zusätzlich deren Blutdruck- und Puls-Parameter erhoben und nach der Tötung die Herzmasse gemessen und auf die Tibialänge relativiert. Ergebnisse: Der von Hinrichs et al. 2011 festgestellte elongierende Effekt von Renin auf Kardiomyozyten konnte bestätigt werden. Unter gleichzeitiger Stimulation mit FCS respektive Endothelin-1 und Renin kam es zur Reduktion eines bekannten zellverbreiternden Effekts von FCS und Endothelin auf die Kardiomyozyten. Diese Reduktion war abhängig von der Reninkonzentration und konnte durch Mannose-6-Phosphat und SB202190 inhibiert werden. Ein verbessernder Effekt der Inkubation mit Renin auf die Parameter der lastfreien Zellverkürzung konnte nicht aufgezeigt werden. Insbesondere im Vergleich der männlichen Tiere haben die reninüberexprimierenden transgenen Ratten signifikant kürzere und schmalere Kardiomyozyten als die vom Blutdruck weitgehend gleichen spontan hypertensiven Ratten. Nur die männlichen transgenen Ratten zeigen auch eine signifikant ausgeprägtere Zellverkürzung auf als die spontan hypertensiven Ratten, jedoch ohne signifikanten Unterschied in der Kontraktions- oder Relaxationsgeschwindigkeit. Diskussion: In vitro zeigt sich ein antihypertropher Effekt von Renin auf Kardiomyozyten. In der Vermittlung dieses Effekts spielen möglicherweise sowohl der M6P/IGFII-Rezeptor als auch der Renin-Rezeptor eine Rolle. Auch die Aktivierung eines intrazellulären RAS nach M6P/IGFII-Rezeptor vermittelter Aufnahme in die Zelle scheint denkbar. In vivo zeigt sich ein ausgeprägter Unterschied zwischen männlichen und weiblichen Tieren. Dies kann einerseits mit der Beeinflussung des RA(A)S durch verschiedene Sexualhomone zusammenhängen, andererseits ist aufgrund seines X-chromosomalen Genlocus auch eine unterschiedliche Expression des PRR denkbar. Die myokardiale Hypertrophie der männlichen transgenen Ratten ist trotz vergleichbarer hämodynamischer Belastung der Herzen weit geringer als die Hypertrophie der spontan hypertensiven Ratten. Weder in vitro durch Stimulation mit Renin noch ex vivo durch erhöhte Reninexpression ließ sich ein wesentlicher positiver Effekt auf die myokardiale Kontraktilität feststellen. Eine Erhöhung der Kontraktions- und Relaxationsgeschwindigkeit bei spontan hypertensiven und transgenen Ratten verglichen mit der Kontrollgruppe aus Sprage-Dawley-Ratten kann auch auf die unterschiedlichen hämodynamischen Bedingungen zurückgeführt werden.Introduction: Myocardial hypertrophy plays a crucial role in the pathophysiology of heart failure. This primarily adaptive mechanism ends in necrosis as well as in apoptotic destruction of myocardial and replacement by connective tissue resulting in a loss of contractile function. A major part in the regulation of myocardial hypertrophy is played by renin-angiotensin (aldosterone) system (RA(A)S). With its hormonal effectors angiotensin II and aldosterone it provokes hypertension and myocardial remodelling. Due to this fact there are many targets for pharmacological therapy of heart failure. In the last decade, several receptors were identified matching the key enzyme of the RA(A)S. By binding to the specific (pro-)renin receptor (PRR) and the mannose-6-phosphat/insulin-like growth factor II receptor (M6P/IGFIIR) the renin can be taken up into the cell and is able to mediate hormonal effects. Also prorenin, the precursor of renin, is able to bind the receptors being internalized and activated to enzymatical activity consecutively. This study examines effects of renin on cardiomyocytes of the rat experiencing prohypertrophic stress. Methods: Cardiac myocytes of male Wistar rats were isolated and taken into short term culture. They were incubated with renin in different concentrations and treated with the known prohypertrophyc stimuli FCS and endothelin-1. After overnight incubation, the length and width of the cells was mesured under the microscope. Furthermore several parameters of cell shortening were collected. To match the results with a rat model ex vivo cardiac myocytes of sprague-dawley rats, spontaneously hypertensive rats (SHR) and transgenic rats TGR(mRen2)27 overexpressing the murine Ren-2 gene were taken into short term culture and subjected to the same mesurements. Before being put to death, the blood pressure and pulse data of the animals were collected. Also the wight of the rats’ hearts in comparrison to the length of their tibia were mesured. Results: The extending effect of renin on cardiac myocytes shown by Hinrichs et al. in 2011 was confirmed. Under stimulation with FCS and endothelin-1 respectively on the one hand and renin on the other hand there was shown a reduction of the brodening effect of FCS and endothelin on cardiac myocytes caused by renin. This reduction was dependent on the concentration of renin and was inhibited by mannose-6-phosphat and SB202190. An ameliorating effect on the parameters of cell shortening could not be shown. Especially regarding male animals the renin overexpressing transgenic rats showed significantly shorter and slimmer cardiac myocytes than the SHR in spite of a comparable blood pressure. Male transgenic rats also showed a greater extend of cell shortening than SHR. There was no significant difference in velocity of contraction and relaxation. Discussion: A antihypertrophic effect of renin on cardiac myocytes is shown in vitro. M6P/IGFIIR as well as PRR may play a role in mediating this effect. Also the activation of an intracellular RAS after M6P/IGFIIR mediated endocytosis of renin or prorenin may be a possible way leading to this effect. There is a distinct difference between male and female animals in vivo. This may be due to the influence of different sexual hormones on RA(A)S but also due to a different expression of the PRR because of ist X-chromosomal locus. Myocardial hypertrophy in male transgenic rats is considerably less developed than in SHR despite comparable hemodynamical stress. A positive effect on the contractility of cardiomyocytes could neither be shown by stimulation by renin in vitro nor by overexpression of renin ex vivo. A higher velocity of contraction and relaxation equally in SHR and transgenic rats compared to sprague-dawley rats may be caused by different hemodynamic conditions

Inducible nitric oxide synthase--time for reappraisal

Inducible nitric oxide synthase (iNOS) is one of three key enzymes generating nitric oxide (NO) from the amino acid L-arginine. iNOS-derived NO plays an important role in numerous physiological and pathophysiological conditions, e.g. blood pressure regulation, inflammation, infection, and the onset and progression of malignant diseases. iNOS has been conjectured both as a marker and a therapeutic target in these situations. iNOS is a mediator of unspecific host defence, central in the clearance of bacterial, viral, fungal and parasitic infections. However, excess production of NO appears to be linked to tissue damage and organ dysfunction, e.g. the hypotensive and vasoplegic state characteristic for septic shock. However, the use of iNOS-inhibitors in septic patients should be performed carefully with regard to the essential functions and properties of NO in blood pressure/blood flow regulation. Considering iNOS-derived NO as a multifactorial transmitter of tumorigenesis and tumor progression, it is tempting to speculate on therapeutical interference with iNOS activity, especially in tumors where metastatic activity, host denfence mechanisms and the level of differentiation seem to be correlated to iNOS expression. It is the aim of this review to provide basic insights into the NOS family of enzymes as well as their regulation. In the second part of the review, we will point out the pivotal roles NOS play in inflammation and neoplastic disease

Inducible nitric oxide synthase (iNOS) in tumor biology: the two sides of the same coin

Inducible nitric oxide synthase (iNOS) is one of three key enzymes generating nitric oxide (NO) from the amino acid l-arginine. iNOS-derived NO plays an important role in numerous physiological (e.g. blood pressure regulation, wound repair and host defence mechanisms) and pathophysiological (inflammation, infection, neoplastic diseases, liver cirrhosis, diabetes) conditions. iNOS is the synthase isoform most commonly associated with malignant disease. Nevertheless, the role of iNOS during tumor development is highly complex, and incompletely understood. Both promoting and deterring actions have been described, presumably depending upon the local concentration of iNOS within the tumor microenvironment. In particular, pivotal effects such as malingnant transformation, angiogenesis, and metastasis are modulated by iNOS. On the other hand, NO derived from macrophages has a potentially cytotoxic/cytostatic effect upon tumor cells. Hence, therapeutical interference with iNOS activity is of considerable interest, especially in tumors where metastatic activity, host defence mechanisms and the level of differentiation seem to be correlated to iNOS expression. This review will aim to summarize the dual actions of iNOS as simultaneous tumor promoter and suppresso

Horsepower of Doctors' Cars Correlates with Cardiovascular Risk and Sedentary Lifestyle but Not with Sexual Dysfunction or Sexual Satisfaction

Background: The horsepower not only of doctors' cars correlates with personal income and social status. However, no clear relationship has previously been described between the horsepower of doctors' cars and cardiovascular health or sexual dysfunction and/or satisfaction. Objective: Cross-sectional online survey to evaluate associations between self-reported horsepower of physicians' cars and health aspects. Methods: Of 1877 physicians from the two University-Hospitals in Austria that were asked to participate in the study, 363 (37.7 ± 8.0 years, 208 (57.3%) men) were included into the final analysis. Results: Physicians that own a car with a stronger engine were significantly older, were more often male, had more often a leading position, had a higher monthly income (all p < 0.001), had a higher scientific output (p = 0.030), and had hypercholesteremia more often (p = 0.009). They also tended to have a higher body mass index (p = 0.088), reported a higher maximum weight in previous years (p = 0.004) and less often reported regular healthy commuting to and from work (p = 0.010). No significant associations were found for self-reported physical fitness, smoking status, and arterial hypertension. In addition, sexual satisfaction and sexual dysfunction were also not related to horsepower in the whole population and the male subgroup. The findings essentially persisted after controlling for age. Conclusion: The horsepower of Austrian physicians' cars correlates with senior position and increased cardiovascular risk. However, our data shows no relationship between sexual dysfunction or lack of sexual satisfaction and the horsepower of doctors' cars

Cisatracurium, but not mivacurium, inhibits survival and axonal growth of neonatal and adult rat peripheral neurons in vitro

Cisatracurium and mivacurium are widely used neuromuscular blocking drugs. Previous reports have indicated growth-inhibitory effects of cisatracurium, but not mivacurium, on two human cell lines in vitro. These effects were ascribed to oxidative stress elicited by acrylate esters formed during cisatracurium breakdown. The aim of the present study was to investigate whether these agents would possibly interfere with the peripheral nervous system. Survival and axonal growth of rat primary neurons obtained from the superior cervical sympathetic ganglion (SCG) or from the adult dorsal root ganglion (DRG) were investigated after treatment with cisatracurium or mivacurium at concentrations from 1 to 10 microM for 24 h. Cisatracurium, but not mivacurium, significantly decreased neuronal survival in a dose-dependent manner, and axonal length was considerably reduced by cisatracurium as compared to controls. It is concluded that high concentrations of cisatracurium are potentially neurotoxi

Anaesthetic implications of Robinow syndrome

Robinow (fetal face) syndrome is a rare inherited multisystem disorder featuring mesomelic or acromesomelic limb shortening, facial and spinal deformities, hypoplastic genitalia, kidney disease and congenital heart defects. We report the anaesthetic management of a patient with Robinow syndrome presenting for elective surgery and review specific issues of interest in the perioperative worku

Parallel SVD-updating using approximate rotations

In this paper a parallel implementation of the SVD-updating algorithm using approximate rotations is presented. In its original form the SVD--updating algorithm had numerical problems if no reorthogonalization steps were applied. Representing the orthogonal matrix V (right singular vectors) using its parameterization in terms of the rotation angles of n(n \Gamma 1)=2 plane rotations these reorthogonalization steps can be avoided during the SVD-updating algorithm [18]. This results in a SVD--updating algorithm where all computations (matrix vector multiplication, QRD--updating, Kogbetliantz&apos;s algorithm) are entirely based on the evaluation and application of orthogonal plane rotations. Therefore, in this form the SVD--updating algorithm is amenable to an implementation using CORDIC--based approximate rotations. Using CORDIC--based approximate rotations the n(n\Gamma1)=2 rotations representing V (as well as all other rotations) are only computed to a certain approximation accuracy (in the basis arctan 2 i). All necessary computations required during the SVD--updating algorithm (exclusively rotations) are executed with the same accuracy, i.e., only r � � w (w: wordlength) elementary orthonormal ��--rotations are used per plane rotation. Simulations show the efficiency of the implementation using CORDIC-based approximate rotations