Komponenten der Bioinkompatibilität der extrakorporalen Hämodialyse des Menschen

Auch heute noch spielt das Phänomen der Bioinkompatibilität von Dialysematerialien eine entscheidende Rolle in der Entstehung von typischen, wenn auch nicht spezifischen, langfristigen Folgeerkrankungen, die aus der chronischen Hämodialyse resultieren. Um das Maß dieser Bioinkompatibilität zu quantifizieren, haben sich einige Parameter besonders gut bewährt. Zur Erfassung gut geeignet ist die Komplementaktivierung im Plasma [5], [9], [42], [70], [109], [110], [112], [144], [193], [197] (sie spiegelt sich insbesondere in der C3a/C3ades Arg- und der C5a/C5ades Arg-Konzentration wider), der Grad der Aktivierung des oxidativen Granulozytenstoffwechsels [51], [79], [104], [109], [134], [135], [143], [181] (Messung z.B. mittels Cytochrom C-Reduktionstest), eine transitorische Leukopenie in der Initialphase der Dialyseprozedur [7], [9], [25], [41], [70], [110], [111], [119], [191], [232], [236], [251] und die Ausschüttung von Interleukinen (z.B. IL-1, IL-6 und TNF) [5], [15], [17], [20], [55], [63], [90], [112], [144], [166], [193], [197], [217]. Bei Urämikern ist die Körpertemperatur niedriger als bei Gesunden [10], [116], [130], [131], [159], [220], [246]. Während der Hämodialyse wird häufig ein Anstieg der Körpertemperatur im Sinne einer febrilen oder hyperthermen Reaktion und ein Anstieg des endogenen Energieumsatzes beobachtet, auch wenn die thermische Energiebilanz im extrakorporalem System gleich Null ist, also weder Wärme zugeführt, noch entzogen wird [47], [131], [139], [140], [141], [142], [145], [146], [159], [167], [168], [169], [170], [205], [209], [145], [146]. Diese Beobachtung wurde auf eine Baroreflex-vermittelte Vasokonstriktion [86], [140] oder auf eine durch Endotoxinfragmente im Dialysat verursachte febrile Reaktion [60], [201], [205] zurückgeführt. Letztlich scheint eine komplexe Summe aus Pyrogenen, Kryogenen und dem Hydratationzustand des Menschen, die Regulation der Körpertemperatur zu bedingen [114], [131]. Für die Steigerung des Energieumsatzes wurde zudem eine vermehrte Atemarbeit verantwortlich gemacht. Diese könnte zwei Ursachen haben: 1. ein aus der Sequestration von Leukozyten in den pulmonalen Kapillaren resultierendes interstitielles Lungenödem könnte die Atemarbeit steigern [140]. 2. der stark erhöhte pCO2 im Bicarbonatdialysat [18] und konsekutiv arteriell könnte ebenfalls einen Anstieg des Atemantriebs bewirken. Der Versuch die Körpertemperatur und den Energieumsatz zur Erfassung der Bioinkompatibilität von Dialysematerialien zu benutzen zeigte, daß beide Parameter nur eingeschränkt dieser Funktion genügen. Sie wirken weniger sensibel als die Erfassung von Komplementsystem, Interleukinen, Leukozytensturz und oxidativem Granulozytenstoffwechsel. Nicht alle Bioinkompatibilitäts-phänomene, vermögen die Körpertemperatur und den Energieumsatz zu beeinflussen. Die vorliegende Untersuchung zeigt eindrücklich, daß weder die Aktivierung des oxidativen Granulozytenstoffwechsels noch die vorübergehende Leukozytopenie ausschließlich membrangebundene Phänomene sind, sondern, wenn auch in geringerem Ausmaß, schon bei einer extrakorporalen Zirkulation ohne Kontakt zu einer Dialysemembran und ohne Dialysatkontakt auftreten. Die Aktivierung des oxidativen Granulozytenstoffwechsels und der Leukozytensturz waren bei alleinigem Kontakt zu einem Dialyseschlauchsystem (Regime A), bei Kontakt zum Schlauchsystem und einer Dialysemembran (Regime B) und bei Kontakt zu Schlauchsystem, Dialysemembran und Dialysatflüssigkeit nachweisbar, jedoch unterschiedlich stark ausgeprägt. Daraus wird deutlich, daß alle Komponenten eines extrakorporalen Dialysekreislaufs einen meßbaren Einfluß auf die Homöostase des Dialysepatienten haben. Verbesserungsbedarf hinsichtlich der Biokompatibilität ist also nicht nur bei den Dialysemembranen vorhanden, sondern auch bei den Schlauchmaterialien und der Dialysatflüssigkeit. Der Kontaminationsgrad des Dialysats mit bakteriellen Zellwandfragmenten wie LPS oder Lipid-A u.a., scheint einen deutlichen Beitrag zu febrilen- und anderen Akutphasereaktionen des Hämodialysepatienten zu leisten. Der Qualität dieser Komponente sollte in der Routine noch mehr Beachtung geschenkt und Kontrollen intensiviert werden. (Die Literaturangaben in den eckigen Klammern beziehen sich auf das Literaturverzeichnis in der Promotion.

Stage progression and neurological symptoms in Trypanosoma brucei rhodesiense sleeping sickness: role of the CNS inflammatory response

Background: Human African trypanosomiasis progresses from an early (hemolymphatic) stage, through CNS invasion to the late (meningoencephalitic) stage. In experimental infections disease progression is associated with neuroinflammatory responses and neurological symptoms, but this concept requires evaluation in African trypanosomiasis patients, where correct diagnosis of the disease stage is of critical therapeutic importance. Methodology/Principal Findings: This was a retrospective study on a cohort of 115 T.b.rhodesiense HAT patients recruited in Eastern Uganda. Paired plasma and CSF samples allowed the measurement of peripheral and CNS immunoglobulin and of CSF cytokine synthesis. Cytokine and immunoglobulin expression were evaluated in relation to disease duration, stage progression and neurological symptoms. Neurological symptoms were not related to stage progression (with the exception of moderate coma). Increases in CNS immunoglobulin, IL-10 and TNF-α synthesis were associated with stage progression and were mirrored by a reduction in TGF-β levels in the CSF. There were no significant associations between CNS immunoglobulin and cytokine production and neurological signs of disease with the exception of moderate coma cases. Within the study group we identified diagnostically early stage cases with no CSF pleocytosis but intrathecal immunoglobulin synthesis and diagnostically late stage cases with marginal CSF pleocytosis and no detectable trypanosomes in the CSF. Conclusions: Our results demonstrate that there is not a direct linkage between stage progression, neurological signs of infection and neuroinflammatory responses in rhodesiense HAT. Neurological signs are observed in both early and late stages, and while intrathecal immunoglobulin synthesis is associated with neurological signs, these are also observed in cases lacking a CNS inflammatory response. While there is an increase in inflammatory cytokine production with stage progression, this is paralleled by increases in CSF IL-10. As stage diagnostics, the CSF immunoglobulins and cytokines studied do not have sufficient sensitivity to be of clinical value

Towards personalised allele-specific CRISPR gene editing to treat autosomal dominant disorders

Abstract CRISPR/Cas9 holds immense potential to treat a range of genetic disorders. Allele-specific gene disruption induced by non-homologous end-joining (NHEJ) DNA repair offers a potential treatment option for autosomal dominant disease. Here, we successfully delivered a plasmid encoding S. pyogenes Cas9 and sgRNA to the corneal epithelium by intrastromal injection and acheived long-term knockdown of a corneal epithelial reporter gene, demonstrating gene disruption via NHEJ in vivo. In addition, we used TGFBI corneal dystrophies as a model of autosomal dominant disease to assess the use of CRISPR/Cas9 in two allele-specific systems, comparing cleavage using a SNP-derived PAM to a guide specific approach. In vitro, cleavage via a SNP-derived PAM was found to confer stringent allele-specific cleavage, while a guide-specific approach lacked the ability to distinguish between the wild-type and mutant alleles. The failings of the guide-specific approach highlights the necessity for meticulous guide design and assessment, as various degrees of allele-specificity are achieved depending on the guide sequence employed. A major concern for the use of CRISPR/Cas9 is its tendency to cleave DNA non-specifically at “off-target” sites. Confirmation that S. pyogenes Cas9 lacks the specificity to discriminate between alleles differing by a single base-pair regardless of the position in the guide is demonstrated

The nuclear receptors of Biomphalaria glabrata and Lottia gigantea: Implications for developing new model organisms

© 2015 Kaur et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are creditedNuclear receptors (NRs) are transcription regulators involved in an array of diverse physiological functions including key roles in endocrine and metabolic function. The aim of this study was to identify nuclear receptors in the fully sequenced genome of the gastropod snail, Biomphalaria glabrata, intermediate host for Schistosoma mansoni and compare these to known vertebrate NRs, with a view to assessing the snail's potential as a invertebrate model organism for endocrine function, both as a prospective new test organism and to elucidate the fundamental genetic and mechanistic causes of disease. For comparative purposes, the genome of a second gastropod, the owl limpet, Lottia gigantea was also investigated for nuclear receptors. Thirty-nine and thirty-three putative NRs were identified from the B. glabrata and L. gigantea genomes respectively, based on the presence of a conserved DNA-binding domain and/or ligand-binding domain. Nuclear receptor transcript expression was confirmed and sequences were subjected to a comparative phylogenetic analysis, which demonstrated that these molluscs have representatives of all the major NR subfamilies (1-6). Many of the identified NRs are conserved between vertebrates and invertebrates, however differences exist, most notably, the absence of receptors of Group 3C, which includes some of the vertebrate endocrine hormone targets. The mollusc genomes also contain NR homologues that are present in insects and nematodes but not in vertebrates, such as Group 1J (HR48/DAF12/HR96). The identification of many shared receptors between humans and molluscs indicates the potential for molluscs as model organisms; however the absence of several steroid hormone receptors indicates snail endocrine systems are fundamentally different.The National Centre for the Replacement, Refinement and Reduction of Animals in Research, Grant Ref:G0900802 to CSJ, LRN, SJ & EJR [www.nc3rs.org.uk]

High-performance work systems and innovation in Vietnamese small firms

This article examines the interplay between high-performance work systems (HPWS) and the innovation of Vietnamese small and medium-sized enterprises (SMEs). Our conceptual model relies on the componential theory of creativity along with HPWS, learning goal orientation (LGO), creativity and innovation to hypothesise both mediation and moderation mechanisms linking such a relationship. Using a sample of 133 SMEs, we find that (1) employee creativity mediates the pathway between HPWS and firm innovation; and (2) LGO moderates the HPWS - employee creativity relationship. Our study casts new light on the theoretical mechanism through which HPWS influence firm innovation and adds to understanding about HPWS within SMEs by bringing employees centre stage

High intensity intermittent games-based activity and adolescents’ cognition: moderating effect of physical fitness

Background: An acute bout of exercise elicits a beneficial effect on subsequent cognitive function in adolescents. The effect of games-based activity, an ecologically valid and attractive exercise model for young people, remains unknown; as does the moderating effect of fitness on the acute exercise-cognition relationship. Therefore, the aim of the present study was to examine the effect of games-based activity on subsequent cognition in adolescents, and the moderating effect of fitness on this relationship. Methods: Following ethical approval, 39 adolescents (12.3 ± 0.7 year) completed an exercise and resting trial in a counterbalanced, randomised crossover design. During familiarisation, participants completed a multi-stage fitness test to predict VO2 peak. The exercise trial consisted of 60-min games-based activity (basketball), during which heart rate was 158 ± 11 beats∙min−1. A battery of cognitive function tests (Stroop test, Sternberg paradigm, trail making and d2 tests) were completed 30-min before, immediately following and 45-min following the basketball. Results: Response times on the complex level of the Stroop test were enhanced both immediately (p = 0.021) and 45-min (p = 0.035) post-exercise, and response times on the five item level of the Sternberg paradigm were enhanced immediately post-exercise (p = 0.023). There were no effects on the time taken to complete the trail making test or any outcome of the d2 test. In particular, response times were enhanced in the fitter adolescents 45-min post-exercise on both levels of the Stroop test (simple, p = 0.005; complex, p = 0.040) and on the three item level of the Sternberg paradigm immediately (p = 0.017) and 45-min (p = 0.008) post-exercise. Conclusions: Games-based activity enhanced executive function and working memory scanning speed in adolescents, an effect particularly evident in fitter adolescents, whilst the high intensity intermittent nature of games-based activity may be too demanding for less fit children

Reply to J.J. Muñoz-Perez et al. Comments on “Confirmation of beach accretion by grain-size trend analysis: Camposoto beach, Cádiz, SWSpain” by E. Poizot et al. (2013) Geo-Marine Letters 33(4)

In a novel finding for a beach environment, Poizot et al. (2013) identified an FB+ trend (sediments becoming finer, better sorted and more positively skewed upshore) on a well-developed swash bar on the upper foreshore of the Camposoto beach of Cádiz in SW Spain. In their Discussion of that paper, Muñoz-Perez et al. (2014) provide some supporting arguments and also report grain-size, beach profile and other data from nearby beaches which differ from those of Poizot and colleagues for Camposoto beach, pointing out that a trend observed on one beach may not apply to a neighbouring beach. However, even though the absolute values differ, the overall trends actually do show the same general behaviour. In our Reply to their comments, we also address some difficulties in comparing granulometric datasets generated by different analytical techniques

Bias and Evolution of the Mutationally Accessible Phenotypic Space in a Developmental System

Genetic and developmental architecture may bias the mutationally available phenotypic spectrum. Although such asymmetries in the introduction of variation may influence possible evolutionary trajectories, we lack quantitative characterization of biases in mutationally inducible phenotypic variation, their genotype-dependence, and their underlying molecular and developmental causes. Here we quantify the mutationally accessible phenotypic spectrum of the vulval developmental system using mutation accumulation (MA) lines derived from four wild isolates of the nematodes Caenorhabditis elegans and C. briggsae. The results confirm that on average, spontaneous mutations degrade developmental precision, with MA lines showing a low, yet consistently increased, proportion of developmental defects and variants. This result indicates strong purifying selection acting to maintain an invariant vulval phenotype. Both developmental system and genotype significantly bias the spectrum of mutationally inducible phenotypic variants. First, irrespective of genotype, there is a developmental bias, such that certain phenotypic variants are commonly induced by MA, while others are very rarely or never induced. Second, we found that both the degree and spectrum of mutationally accessible phenotypic variation are genotype-dependent. Overall, C. briggsae MA lines exhibited a two-fold higher decline in precision than the C. elegans MA lines. Moreover, the propensity to generate specific developmental variants depended on the genetic background. We show that such genotype-specific developmental biases are likely due to cryptic quantitative variation in activities of underlying molecular cascades. This analysis allowed us to identify the mutationally most sensitive elements of the vulval developmental system, which may indicate axes of potential evolutionary variation. Consistent with this scenario, we found that evolutionary trends in the vulval system concern the phenotypic characters that are most easily affected by mutation. This study provides an empirical assessment of developmental bias and the evolution of mutationally accessible phenotypes and supports the notion that such bias may influence the directions of evolutionary change

Genome-wide binding of the CRISPR endonuclease Cas9 in mammalian cells

Bacterial type II CRISPR-Cas9 systems have been widely adapted for RNA-guided genome editing and transcription regulation in eukaryotic cells, yet their in vivo target specificity is poorly understood. Here we mapped genome-wide binding sites of a catalytically inactive Cas9 (dCas9) from Streptococcus pyogenes loaded with single guide RNAs (sgRNAs) in mouse embryonic stem cells (mESCs). Each of the four sgRNAs we tested targets dCas9 to between tens and thousands of genomic sites, frequently characterized by a 5-nucleotide seed region in the sgRNA and an NGG protospacer adjacent motif (PAM). Chromatin inaccessibility decreases dCas9 binding to other sites with matching seed sequences; thus 70% of off-target sites are associated with genes. Targeted sequencing of 295 dCas9 binding sites in mESCs transfected with catalytically active Cas9 identified only one site mutated above background levels. We propose a two-state model for Cas9 binding and cleavage, in which a seed match triggers binding but extensive pairing with target DNA is required for cleavage.National Institutes of Health (U.S.) (Grant RO1-GM34277)National Institutes of Health (U.S.) (Grant R01-CA133404)National Cancer Institute (U.S.) (Grant PO1-CA42063)National Cancer Institute (U.S.) (Cancer Center Support (Core) Grant P30-CA14051)National Institutes of Health (U.S.) (Director's Pioneer Award 1DP1-MH100706)Damon Runyon Cancer Research FoundationKinship Foundation. Searle Scholars ProgramSimons Foundatio