The influence of normobaric hyperoxia on hepatic oxygenation--experience with an animal model

We investigated the effect of a ventilation with an FiO2 of 1.0 on arterial and hepatic venous oxygenation in 23 G?ttingen minipigs. Under balanced anaesthesia (isoflurane/fentanyl), a fibreoptic catheter was placed into a hepatic vein. The correct position of the tip of the catheter was controlled manually after laparotomy. After measurement of baseline values (arterial and hepatic blood gases, ShvO2), in 13 minipigs normoventilation with an FiO2 of 1.0 was performed for 15 minutes. Thereafter, ventilation was continued with an FiO2 of 0.4. In the control group (n = 10), the animals were oxygenated with an FiO2 of 0.4 permanently. The changes due to hyperoxia were measured in hepatic venous oxygen saturation (ShvbgaO2: from 81.2 +/- 1.43% to 87.5 +/- 1.77%, ShvoximO2: from 82.6 +/- 1.14% to 90.5 +/- 0.90%), arterial (from 217.5 +/- 5.0 mmHg to 467.2 +/- 22.0 mmHg) and hepatic venous (from 51.8 +/- 2.0 mmHg) oxygen partial pressure. We found a correlation between hepatic venous oxygen partial pressure und ShvbgaO2 in the blood (r = 0.84, p < 0.001) and between ShvO2 (ShvbgaO2/ShvoximO2), which was either measured directly in the blood or by a fibreoptic catheter (r = 0.6, p < 0.001). Whereas the increase in ShvO2 during hyperoxia may be a result of increased arterial supply, the decrease in ShvO2 after the end of hyperoxia below baseline values needs further investigations. The continuous fibreoptic measurement of ShvoximO2, also under hyperoxic conditions is a valuable parameter for the monitoring of hepatic venous oxygenation.     

In vivo 3D MRI staining of the mouse hippocampal system using intracerebral injection of MnCl2

The morphology and function of the hippocampal system of C57BL/6J mice (n = 8) was studied in vivo using T1-weighted 3D magnetic resonance imaging (MRI) (117 microm isotropic resolution) after bilateral injection of MnCl(2) (0.25 microl, 5 or 200 mM) into the posterior hippocampal formation. The neuronal uptake of the T1-shortening Mn(2+) ions resulted in a pronounced MRI signal enhancement within the CA3 subfield and dentate gyrus with milder increases in CA1 and subiculum. This finding is in line with differences in the excitability of hippocampal neurons previously reported using electrophysiologic recordings. The subsequent axonal transport of Mn(2+) highlighted the principal extrinsic projections from the posterior hippocampal formation via the fimbria and the precommissural fornix to the dorsal part of the lateral septal nucleus. A strong MRI signal enhancement was also observed in the ventral hippocampal commissure. A time-course analysis revealed unsaturated conditions of Mn(2+) accumulation at about 2 h after injection and optimal contrast-to-noise ratios at about 6 h after injection. The present results using Mn(2+)-enhanced 3D MRI open new ways for studying the role of the hippocampal system in specific aspects of learning and memory in normal and mutant mice.     

Combined expression of tau and the Harlequin mouse mutation leads to increased mitochondrial dysfunction, tau pathology and neurodegeneration

Mitochondrial dysfunction and oxidative stress play an important role in ageing and have been implicated in several age-related neurodegenerative conditions including Alzheimer's disease (AD) and other tauopathies characterized by the presence of intracellular accumulations of the hyperphosphorylated microtubule-associated protein tau. To study the interaction between mitochondrial dysfunction and tau pathology in vivo, we generated a novel mouse model by crossbreeding two existing lines: the Harlequin (Hq) mutant mice which suffer from mitochondrial dysfunction and oxidative stress due to a lack of the mitochondrial apoptosis-inducing factor (AIF), and the P301L tau transgenic mice, a mouse model of human tau pathology.Combined expression of the Hq mouse mutation and the tau transgene in the Tau/Hq double mutant mice led to an increase in tau pathology and apoptotic neurodegeneration when compared to single expression of the two mutations. Neurodegeneration was most prominent in the dentate gyrus and was significantly increased in the cerebellum leading to aggravated motor deficits. Functional activity measurements of the mitochondrial respiratory chain (MRC) in the Tau/Hq mice revealed early decreased activities of multiple MRC complexes and depleted ATP levels which preceded neurodegeneration and elevated oxidative stress markers. These results suggest an age-dependent mutual reinforcement of the tau pathology and mitochondrial dysfunction in vivo, which may contribute to neurodegeneration in patients suffering from AD and other age-related tauopathies.     

Streptococcus pneumoniae Autolysis Prevents Phagocytosis and Production of Phagocyte-Activating Cytokines

Streptococcus pneumoniae is a major pathogen in humans. The pathogenicity of this organism is related to its many virulence factors, the most important of which is the thick pneumococcal capsule that minimizes phagocytosis. Another virulence-associated trait is the tendency of this bacterium to undergo autolysis in stationary phase through activation of the cell wall-bound amidase LytA, which breaks down peptidoglycan. The exact function of autolysis in pneumococcal pathogenesis is, however, unclear. Here, we show the selective and specific inefficiency of wild-type S. pneumoniae for inducing production of phagocyte-activating cytokines in human peripheral blood mononuclear cells (PBMC). Indeed, clinical pneumococcal strains induced production of 30-fold less tumor necrosis factor (TNF), 15-fold less gamma interferon (IFN-γ), and only negligible amounts of interleukin-12 (IL-12) compared with other closely related Streptococcus species, whereas the levels of induction of IL-6, IL-8, and IL-10 production were similar. If pneumococcal LytA was inactivated by mutation or by culture in a medium containing excess choline, the pneumococci induced production of significantly more TNF, IFN-γ, and IL-12 in PBMC, whereas the production of IL-6, IL-8, and IL-10 was unaffected. Further, adding autolyzed pneumococci to intact bacteria inhibited production of TNF, IFN-γ, and IL-12 in a dose-dependent manner but did not inhibit production of IL-6, IL-8, and IL-10 in response to the intact bacteria. Fragments from autolyzed bacteria inhibited phagocytosis of intact bacteria and reduced the in vitro elimination of pneumococci from human blood. Our results suggest that fragments generated by autolysis of bacteria with reduced viability interfere with phagocyte-mediated elimination of live pneumococci.The pneumococcus Streptococcus pneumoniae is a leading cause of community-acquired pneumonia, meningitis, otitis media, and sinusitis and is a common cause of infection-related mortality in children and elderly people (28, 37).There is a large number of streptococcal species whose taxonomic classification is debated (14, 31). A number of streptococci, including alpha-hemolytic and nonhemolytic variants, constitute the viridans group, which can be further subdivided into the mitis, sanguinis, anginosus, salivarius, and mutans groups based on biochemical tests (14). Phenotypic and genetic tests consistently show that S. pneumoniae is closely related to and may be placed in the mitis subgroup (14, 30). Although the other members of the mitis group can cause sepsis and endocarditis (53), they are considerably less virulent than S. pneumoniae.Pneumococci are considered strictly extracellular pathogens, whose elimination depends on ingestion and killing by phagocytes (i.e., alveolar and tissue-resident macrophages and neutrophils recruited during the inflammatory process). Accordingly, an important determinant of pneumococcal pathogenicity is the thick, hydrophilic polysaccharide capsule, which impedes elimination by phagocytes in the absence of capsule-specific antibodies.The ability of phagocytes to kill microbes is augmented by the phagocyte-activating cytokines gamma interferon (IFN-γ) and tumor necrosis factor (TNF), which boost the bactericidal machinery and enhance killing and digestion of bacteria present within the phagosome (4, 39, 47). TNF is produced by monocytes/macrophages and activated T cells, while IFN-γ is produced by NK cells and T cells in response to interleukin-12 (IL-12) from macrophages. Thus, production of TNF, IFN-γ, and IL-12 is necessary for host defense against intracellular bacteria (8, 11, 21, 34, 48). More recently, these phagocyte-activating cytokines have also been shown to be essential for controlling extracellular gram-positive bacteria, including S. pneumoniae (36, 42, 50, 52, 54). Thus, a patient with an IL-12 deficiency was shown to suffer from recurrent episodes of pneumococcal infection (20). Phagocyte activation by TNF and/or IFN-γ might be required for decomposition of the thick, sturdy peptidoglycan (PG) layer of gram-positive bacteria after phagocytosis, while gram-negative bacteria may be more easily digested. Thus, human leukocytes produce more TNF, IFN-γ, and IL-12 when they are stimulated with gram-positive bacteria than when they are stimulated with gram-negative bacteria (23, 24).A peculiar property of S. pneumoniae is its tendency to undergo autolysis when it reaches the stationary phase of growth. This process is mediated by enzymes called autolysins (ALs), which, when activated, degrade cell wall PG. The major AL is an N-acetyl-muramyl-l-alanine amidase called LytA (27). Other pneumococcal ALs include LytB and LytC, which are believed to be involved mainly in modification of the cell wall during growth and division (16, 17). ALs are anchored to the cell wall via interactions with choline moieties on teichoic acid and lipoteichoic acid (LTA). Choline is necessary for pneumococcal growth, but culture in the presence of high concentrations of choline renders the bacteria incapable of undergoing autolysis (6, 19).Studies with mice have shown that S. pneumoniae with mutated LytA is less virulent than wild-type pneumococci (2, 7, 25). The reason for this is not clear, but two main hypotheses have been put forward. First, autolysis promotes the release of the intracellular toxin pneumolysin (Ply) (5, 33). Ply is an important determinant of virulence (3, 41) and interferes with several defense systems, including inhibition of ciliary beating (15), complement activation (38), and induction of intracellular oxygen radical production (33). Second, cell wall degradation products, such as soluble PG fragments and LTA released upon autolysis, have been suggested to augment the inflammatory response (9, 10, 44, 49).Here we examine a third possibility, that autolysis interferes with the generation of phagocyte-activating cytokines. We have previously shown that intact gram-positive bacteria provide a very efficient stimulus for IL-12 production by human monocytes, regardless of whether they are dead or alive (1, 23, 24), but that decomposed bacteria are inactive in this process and soluble components of the gram-positive cell wall, such as PG and LTA, even downregulate the production of IL-12 in response to intact bacteria in a dose-dependent manner (1). These observations led us to speculate that autolysis may promote virulence by generating bacterial cell wall fragments that block IL-12 production and thereby reduce IFN-γ production and phagocyte activation. Indeed, our data demonstrate that AL-mediated disintegration of pneumococci inhibits production of IFN-γ and also TNF in response to intact bacteria. Further, the presence of autolyzed bacteria reduced elimination of live pneumococci by blood cells in vitro.     

Functional protein expression of multiple sodium channel α- and β-subunit isoforms in neonatal cardiomyocytes

Voltage-gated sodium channels are composed of pore-forming α- and auxiliary β-subunits and are responsible for the rapid depolarization of cardiac action potentials. Recent evidence indicates that neuronal tetrodotoxin (TTX) sensitive sodium channel α-subunits are expressed in the heart in addition to the predominant cardiac TTX-resistant Na_v1.5 sodium channel α-subunit. These TTX-sensitive isoforms are preferentially localized in the transverse tubules of rodents. Since neonatal cardiomyocytes have yet to develop transverse tubules, we determined the complement of sodium channel subunits expressed in these cells. Neonatal rat ventricular cardiomyocytes were stained with antibodies specific for individual isoforms of sodium channel α- and β-subunits. α-actinin, a component of the z-line, was used as an intracellular marker of sarcomere boundaries. TTX-sensitive sodium channel α-subunit isoforms Na_v1.1, Na_v1.2, Na_v1.3, Na_v1.4 and Na_v1.6 were detected in neonatal rat heart but at levels reduced compared to the predominant cardiac α-subunit isoform, Na_v1.5. Each of the β-subunit isoforms (β1-β4) was also expressed in neonatal cardiac cells. In contrast to adult cardiomyocytes, the α-subunits are distributed in punctate clusters across the membrane surface of neonatal cardiomyocytes; no isoform-specific subcellular localization is observed. Voltage clamp recordings in the absence and presence of 20 nM TTX provided functional evidence for the presence of TTX-sensitive sodium current in neonatal ventricular myocardium which represents between 20 and 30% of the current, depending on membrane potential and experimental conditions. Thus, as in the adult heart, a range of sodium channel α-subunits are expressed in neonatal myocytes in addition to the predominant TTX-resistant Na_v1.5 α-subunit and they contribute to the total sodium current.     

How sharp is the short QuickDASH? A refined content and validity analysis of the short form of the disabilities of the shoulder, arm and hand questionnaire in the strata of symptoms and function and specific joint conditions

Purpose  

To assess and compare content, validity, and specificity of the QuickDASH (Disability of the arm, shoulder and hand questionnaire) as compared to the full-length DASH and other instruments to give a recommendation for its use depending on a specific clinical situation.     

Conditional neuronal nitric oxide synthase overexpression impairs myocardial contractility

The role of the neuronal NO synthase (nNOS or NOS1) enzyme in the control of cardiac function still remains unclear. Results from nNOS(-/-) mice or from pharmacological inhibition of nNOS are contradictory and do not pay tribute to the fact that probably spatial confinement of the nNOS enzyme is of major importance. We hypothesize that the close proximity of nNOS and certain effector molecules like L-type Ca(2+)-channels has an impact on myocardial contractility. To test this, we generated a new transgenic mouse model allowing conditional, myocardial specific nNOS overexpression. Western blot analysis of transgenic nNOS overexpression showed a 6-fold increase in nNOS protein expression compared with noninduced littermates (n=12; P<0.01). Measuring of total NOS activity by conversion of [(3)H]-l-arginine to [(3)H]-l-citrulline showed a 30% increase in nNOS overexpressing mice (n=18; P<0.05). After a 2 week induction, nNOS overexpression mice showed reduced myocardial contractility. In vivo examinations of the nNOS overexpressing mice revealed a 17+/-3% decrease of +dp/dt(max) compared with noninduced mice (P<0.05). Likewise, ejection fraction was reduced significantly (42% versus 65%; n=15; P<0.05). Interestingly, coimmunoprecipitation experiments indicated interaction of nNOS with SR Ca(2+)ATPase and additionally with L-type Ca(2+)- channels in nNOS overexpressing animals. Accordingly, in adult isolated cardiac myocytes, I(Ca,L) density was significantly decreased in the nNOS overexpressing cells. Intracellular Ca(2+)-transients and fractional shortening in cardiomyocytes were also clearly impaired in nNOS overexpressing mice versus noninduced littermates. In conclusion, conditional myocardial specific overexpression of nNOS in a transgenic animal model reduced myocardial contractility. We suggest that nNOS might suppress the function of L-type Ca(2+)-channels and in turn reduces Ca(2+)-transients which accounts for the negative inotropic effect.     

Successful interventions on an organisational level to reduce violence and coercive interventions in in-patients with adjustment disorders and personality disorders

Background

Self-directed and other violence as well as subsequent coercive interventions occur in a substantial proportion of patients with personality disorders during in-patient treatment. Different strategies may be required to reduce coercive interventions for patients of different diagnostic groups.

Methods

We specialised one of our acute admission wards in the treatment of personality disorders and adjustment disorders (ICD-10 F4 and F6). Patients are not transferred to other acute wards in case of suicidal or violent behaviour. Violent behaviour and coercive interventions such as seclusion or restraint were recorded in the same way as in the rest of the hospital. We recorded the percentage of subjects affected by diagnostic group and average length of an intervention in the year before and after the change in organisational structure.

Results

The total number of coercive interventions decreased by 85% both among patients with an F4 and those with an F6 primary diagnosis. Violent behaviours decreased by about 50%, the proportion of involuntary committed patients decreased by 70%.

Conclusion

The organisational change turned out to be highly effective without any additional cost of personnel or other resources.