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Background:
Oral anticoagulant therapy (OAT) potently prevents strokes in patients with atrial fibrillation. Vitamin K antagonists (VKA) have been the standard of care for long-term OAT for decades, but non-VKA oral anticoagulants (NOAC) have recently been approved for this indication, and raised many questions, among them their influence on medication adherence. We assessed adherence to VKA and NOAC in secondary stroke prevention.
Methods:
All patients treated from October 2011 to September 2012 for ischemic stroke or transient ischemic attack with a subsequent indication for OAT, at three academic hospitals were entered into a prospective registry, and baseline data and antithrombotic treatment at discharge were recorded. At the 1-year follow-up, we assessed the adherence to different OAT strategies and patients' adherence to their respective OAT. We noted OAT changes, reasons to change treatment, and factors that influence persistence to the prescribed OAT.
Results:
In patients discharged on OAT, we achieved a fatality corrected response rate of 73.3% (n=209). A total of 92% of these patients received OAT at the 1-year follow-up. We observed good adherence to both VKA and NOAC (VKA, 80.9%; NOAC, 74.8%; P=0.243) with a statistically nonsignificant tendency toward a weaker adherence to dabigatran. Disability at 1-year follow-up was an independent predictor of lower adherence to any OAT after multivariate analysis, whereas the choice of OAT did not have a relevant influence.
Conclusion:
One-year adherence to OAT after stroke is strong (>90%) and patients who switch therapy most commonly switch toward another OAT. The 1-year adherence rates to VKA and NOAC in secondary stroke prevention do not differ significantly between both therapeutic strategies.
In today’s social online world there is a variety of interaction and participatory possibilities which enable web users to actively produce content themselves.
This user-generated content is omnipresent in the web and there is growing evidence that it is used to select or evaluate professionally created online information.
The present study investigated how this surrounding content affects online advertising by drawing from social influence theory. Specifically, it was assumed that
web users sharing an interpersonal relationship (interpersonal influence) and/or a group membership (collective influence) with authors of user-generated content
which appears next to advertising on the web page are more strongly influenced in their response to the advertising than unrelated users. These assumptions were
tested in a 2 × 2 between-subject experiment with 118 students who were exposed to four different Facebook profiles that differed in terms of interpersonal
connection to the source (existent/non-existent) and collective connection to the source (existent/non-existent). The results show a significant impact in the case
of collective influence, but not in the case of interpersonal influence. The underlying mechanisms of this effect and implications of the results for online advertising
are discussed.
To investigate the usefulness of pain-related evoked potentials (PREP) elicited by electrical stimulation for the identification of small fiber involvement in patients with mixed fiber neuropathy (MFN). Eleven MFN patients with clinical signs of large fiber impairment and neuropathic pain and ten healthy controls underwent clinical and electrophysiological evaluation. Small fiber function, electrical conductivity and morphology were examined by quantitative sensory testing (QST), PREP, and skin punch biopsy. MFN was diagnosed following clinical and electrophysiological examination (chronic inflammatory demyelinating neuropathy: n = 6; vasculitic neuropathy: n = 3; chronic axonal neuropathy: n = 2). The majority of patients with MFN characterized their pain by descriptors that mainly represent C-fiber-mediated pain. In QST, patients displayed elevated cold, warm, mechanical, and vibration detection thresholds and cold pain thresholds indicative of MFN. PREP amplitudes in patients correlated with cold (p < 0.05) and warm detection thresholds (p < 0.05). Burning pain and the presence of par-/dysesthesias correlated negatively with PREP amplitudes (p < 0.05). PREP amplitudes correlating with cold and warm detection thresholds, burning pain, and par-/dysesthesias support employing PREP amplitudes as an additional tool in conjunction with QST for detecting small fiber impairment in patients with MFN.
Background
To use combinatorial epitope mapping ("fingerprinting") of the antibody response to identify targets of the humoral immune response in patients with transitional cell carcinoma (TCC) of the bladder.
Methods
A combinatorial random peptide library was screened on the circulating pool of immunoglobulins purified from an index patient with a high risk TCC (pTa high grade plus carcinoma in situ) to identify corresponding target antigens. A patient cohort was investigated for antibody titers against ubiquitin.
Results
We selected, isolated, and validated an immunogenic peptide motif from ubiquitin as a dominant epitope of the humoral response. Patients with TCC had significantly higher antibody titers against ubiquitin than healthy donors (p<0.007), prostate cancer patients (p<0.0007), and all patients without TCC taken together (p<0.0001). Titers from superficial tumors were not significantly different from muscle invasive tumors (p = 0.0929). For antibody response against ubiquitin, sensitivity for detection of TCC was 0.44, specificity 0.96, positive predictive value 0.96 and negative predictive value 0.41. No significant titer changes were observed during the standard BCG induction immunotherapy.
Conclusions
This is the first report to demonstrate an anti-ubiquitin antibody response in patients with TCC. Although sensitivity of antibody production was low, a high specificity and positive predictive value make ubiquitin an interesting candidate for further diagnostic and possibly immune modulating studies.
Brain-computer interfaces (BCIs) can serve as muscle independent communication aids. Persons, who are unable to control their eye muscles (e.g., in the completely locked-in state) or have severe visual impairments for other reasons, need BCI systems that do not rely on the visual modality. For this reason, BCIs that employ auditory stimuli were suggested. In this study, a multiclass BCI spelling system was implemented that uses animal voices with directional cues to code rows and columns of a letter matrix. To reveal possible training effects with the system, 11 healthy participants performed spelling tasks on 2 consecutive days. In a second step, the system was tested by a participant with amyotrophic lateral sclerosis (ALS) in two sessions. In the first session, healthy participants spelled with an average accuracy of 76% (3.29 bits/min) that increased to 90% (4.23 bits/min) on the second day. Spelling accuracy by the participant with ALS was 20% in the first and 47% in the second session. The results indicate a strong training effect for both the healthy participants and the participant with ALS. While healthy participants reached high accuracies in the first session and second session, accuracies for the participant with ALS were not sufficient for satisfactory communication in both sessions. More training sessions might be needed to improve spelling accuracies. The study demonstrated the feasibility of the auditory BCI with healthy users and stresses the importance of training with auditory multiclass BCIs, especially for potential end-users of BCI with disease.
Introduction:
Individuals carrying pathogenic mutations in the BRCA1 and BRCA2 genes have a high lifetime risk of breast cancer. BRCA1 and BRCA2 are involved in DNA double-strand break repair, DNA alterations that can be caused by exposure to reactive oxygen species, a main source of which are mitochondria. Mitochondrial genome variations affect electron transport chain efficiency and reactive oxygen species production. Individuals with different mitochondrial haplogroups differ in their metabolism and sensitivity to oxidative stress. Variability in mitochondrial genetic background can alter reactive oxygen species production, leading to cancer risk. In the present study, we tested the hypothesis that mitochondrial haplogroups modify breast cancer risk in BRCA1/2 mutation carriers.
Methods:
We genotyped 22,214 (11,421 affected, 10,793 unaffected) mutation carriers belonging to the Consortium of Investigators of Modifiers of BRCA1/2 for 129 mitochondrial polymorphisms using the iCOGS array. Haplogroup inference and association detection were performed using a phylogenetic approach. ALTree was applied to explore the reference mitochondrial evolutionary tree and detect subclades enriched in affected or unaffected individuals.
Results:
We discovered that subclade T1a1 was depleted in affected BRCA2 mutation carriers compared with the rest of clade T (hazard ratio (HR) = 0.55; 95% confidence interval (CI), 0.34 to 0.88; P = 0.01). Compared with the most frequent haplogroup in the general population (that is, H and T clades), the T1a1 haplogroup has a HR of 0.62 (95% CI, 0.40 to 0.95; P = 0.03). We also identified three potential susceptibility loci, including G13708A/rs28359178, which has demonstrated an inverse association with familial breast cancer risk.
Conclusions:
This study illustrates how original approaches such as the phylogeny-based method we used can empower classical molecular epidemiological studies aimed at identifying association or risk modification effects.
An Overview of the Regional Experiments for Land-atmosphere Exchanges 2012 (REFLEX 2012) Campaign
(2015)
The REFLEX 2012 campaign was initiated as part of a training course on the organization of an airborne campaign to support advancement of the understanding of land-atmosphere interaction processes. This article describes the campaign, its objectives and observations, remote as well as in situ. The observations took place at the experimental Las Tiesas farm in an agricultural area in the south of Spain. During the period of ten days, measurements were made to capture the main processes controlling the local and regional land-atmosphere exchanges. Apart from multi-temporal, multi-directional and multi-spatial space-borne and airborne observations, measurements of the local meteorology, energy fluxes, soil temperature profiles, soil moisture profiles, surface temperature, canopy structure as well as leaf-level measurements were carried out. Additional thermo-dynamical monitoring took place at selected sites. After presenting the different types of measurements, some examples are given to illustrate the potential of the observations made.
The gene encoding the LIM and SH3 domain protein (LASP1) was cloned two decades ago from a cDNA library of breast cancer metastases. As the first protein of a class comprising one N-terminal LIM and one C-terminal SH3 domain, LASP1 founded a new LIM-protein subfamily of the nebulin group. Since its discovery LASP1 proved to be an extremely versatile protein because of its exceptional structure allowing interaction with various binding partners, its ubiquitous expression in normal tissues, albeit with distinct expression patterns, and its ability to transmit signals from the cytoplasm into the nucleus. As a result, LASP1 plays key roles in cell structure, physiological processes, and cell signaling. Furthermore, LASP1 overexpression contributes to cancer aggressiveness hinting to a potential value of LASP1 as a cancer biomarker. In this review we summarize published data on structure, regulation, function, and expression pattern of LASP1, with a focus on its role in human cancer and as a biomarker protein. In addition, we provide a comprehensive transcriptome analysis of published microarrays (n=2,780) that illustrates the expression profile of LASP1 in normal tissues and its overexpression in a broad range of human cancer entities.
Analysis of a multi-component multi-stage malaria vaccine candidate—tackling the cocktail challenge
(2015)
Combining key antigens from the different stages of the P. falciparum life cycle in the context of a multi-stage-specific cocktail offers a promising approach towards the development of a malaria vaccine ideally capable of preventing initial infection, the clinical manifestation as well as the transmission of the disease. To investigate the potential of such an approach we combined proteins and domains (11 in total) from the pre-erythrocytic, blood and sexual stages of P. falciparum into a cocktail of four different components recombinantly produced in plants. After immunization of rabbits we determined the domain-specific antibody titers as well as component-specific antibody concentrations and correlated them with stage specific in vitro efficacy. Using purified rabbit immune IgG we observed strong inhibition in functional in vitro assays addressing the pre-erythrocytic (up to 80%), blood (up to 90%) and sexual parasite stages (100%). Based on the component-specific antibody concentrations we calculated the IC50 values for the pre-erythrocytic stage (17–25 μg/ml), the blood stage (40–60 μg/ml) and the sexual stage (1.75 μg/ml). While the results underline the feasibility of a multi-stage vaccine cocktail, the analysis of component-specific efficacy indicates significant differences in IC50 requirements for stage-specific antibody concentrations providing valuable insights into this complex scenario and will thereby improve future approaches towards malaria vaccine cocktail development regarding the selection of suitable antigens and the ratios of components, to fine tune overall and stage-specific efficacy.
This review outlines the most frequently used rodent stroke models and discusses their strengths and shortcomings. Mimicking all aspects of human stroke in one animal model is not feasible because ischemic stroke in humans is a heterogeneous disorder with a complex pathophysiology. The transient or permanent middle cerebral artery occlusion (MCAo) model is one of the models that most closely simulate human ischemic stroke. Furthermore, this model is characterized by reliable and well-reproducible infarcts. Therefore, the MCAo model has been involved in the majority of studies that address pathophysiological processes or neuroprotective agents. Another model uses thromboembolic clots and thus is more convenient for investigating thrombolytic agents and pathophysiological processes after thrombolysis. However, for many reasons, preclinical stroke research has a low translational success rate. One factor might be the choice of stroke model. Whereas the therapeutic responsiveness of permanent focal stroke in humans declines significantly within 3 hours after stroke onset, the therapeutic window in animal models with prompt reperfusion is up to 12 hours, resulting in a much longer action time of the investigated agent. Another major problem of animal stroke models is that studies are mostly conducted in young animals without any comorbidity. These models differ from human stroke, which particularly affects elderly people who have various cerebrovascular risk factors. Choosing the most appropriate stroke model and optimizing the study design of preclinical trials might increase the translational potential of animal stroke models.
Bornyl caffeate (1) was previously isolated by us from Valeriana (V.) wallichii rhizomes and identified as an anti-leishmanial substance. Here, we screened a small compound library of synthesized derivatives 1–30 for activity against schistosomula of Schistosoma (S.) mansoni. Compound 1 did not show any anti-schistosomal activity. However, strong phenotypic changes, including the formation of vacuoles, degeneration and death were observed after in vitro treatment with compounds 23 (thymyl cinnamate) and 27 (eugenyl cinnamate). Electron microscopy analysis of the induced vacuoles in the dying parasites suggests that 23 and 27 interfere with autophagy.
Peptides derived from human and bovine lactoferricin were designed, synthesized, purified, and characterized using RP-HPLC and MALDI-TOF-MS. Specific changes in the sequences were designed as (i) the incorporation of unnatural amino acids in the sequence, the (ii) reduction or (iii) elongation of the peptide chain length, and (iv) synthesis of molecules with different number of branches containing the same sequence. For each peptide, the antibacterial activity against Escherichia coli ATCC 25922 and Enterococcus faecalis ATCC 29212 was evaluated. Our results showed that Peptides I.2 (RWQWRWQWR) and I.4 ((RRWQWR)\(_{4}\)K\(_{2}\)Ahx\(_{2}\)C\(_{2}\)) exhibit bigger or similar activity against E. coli (MIC 4-33 μM) and E. faecalis (MIC 10-33 μM) when they were compared with lactoferricin protein (LF) and some of its derivate peptides as II.1 (FKCRRWQWRMKKLGA) and IV.1 (FKCRRWQWRMKKLGAPSITCVRRAE). It should be pointed out that Peptides I.2 and I.4, containing the RWQWR motif, are short and easy to synthesize; our results demonstrate that it is possible to design and obtain synthetic peptides that exhibit enhanced antibacterial activity using a methodology that is fast and low-cost and that allows obtaining products with a high degree of purity and high yield.
The chloroform extract of Valeriana wallichii (V. wallichii) rhizomes was investigated to elucidate the structures responsible for reported antileishmanial activity. Besides bornyl caffeate (1, already been reported by us previously), bioassay-guided fractionation resulted in two additional cinnamic acid derivatives 2–3 with moderate leishmanicidal activity. The structure of a novel nepetolactone derivative 4 having a cinnamic acid moiety was elucidated by means of spectral analysis. To the best of our knowledge villoside aglycone (5) was isolated from this plant for the first time. The bioassay-guided fractionation yielded two new (compounds 6–7) and two known valtrates (compounds 8–9) with leishmanicidal potential against Leishmania major (L. major) promastigotes. In addition, β-bisabolol (10), α-kessyl alcohol (11), valeranone (12), bornyl isovalerate (13) and linarin-2-O-methylbutyrate (14) were identified. This is the first report on the isolation of 4'-demethylpodophyllotoxin (15), podophyllotoxin (16) and pinoresinol (17) in V. wallichii. In total thirteen known and four new compounds were identified from the extract and their cytotoxic and antileishmanial properties were evaluated.
In classical conditioning, an initially neutral stimulus (conditioned stimulus, CS) becomes associated with a biologically salient event (unconditioned stimulus, US), which might be pain (aversive conditioning) or food (appetitive conditioning). After a few associations, the CS is able to initiate either defensive or consummatory responses, respectively. Contrary to aversive conditioning, appetitive conditioning is rarely investigated in humans, although its importance for normal and pathological behaviors (e.g., obesity, addiction) is undeniable. The present study intents to translate animal findings on appetitive conditioning to humans using food as an US. Thirty-three participants were investigated between 8 and 10 am without breakfast in order to assure that they felt hungry. During two acquisition phases, one geometrical shape (avCS+) predicted an aversive US (painful electric shock), another shape (appCS+) predicted an appetitive US (chocolate or salty pretzel according to the participants' preference), and a third shape (CS) predicted neither US. In a extinction phase, these three shapes plus a novel shape (NEW) were presented again without US delivery. Valence and arousal ratings as well as startle and skin conductance (SCR) responses were collected as learning indices. We found successful aversive and appetitive conditioning. On the one hand, the avCS+ was rated as more negative and more arousing than the CS and induced startle potentiation and enhanced SCR. On the other hand, the appCS+ was rated more positive than the CS and induced startle attenuation and larger SCR. In summary, we successfully confirmed animal findings in (hungry) humans by demonstrating appetitive learning and normal aversive learning.
Human upcyte\(^{®}\) hepatocytes are proliferating hepatocytes that retain many characteristics of primary human hepatocytes. We conducted a comprehensive evaluation of the application of second-generation upcyte\(^{®}\) hepatocytes from four donors for inhibition and induction assays using a selection of reference inhibitors and inducers. CYP1A2, CYP2B6, CYP2C9, and CYP3A4 were reproducibly inhibited in a concentration-dependent manner and the calculated IC\(_{50}\) values for each compound correctly classified them as potent inhibitors. Upcyte\(^{®}\) hepatocytes were responsive to prototypical CYP1A2, CYP2B6, CYP2C9, and CYP3A4 inducers, confirming that they have functional AhR-, CAR-, and PXR-mediated CYP regulation. A panel of 11 inducers classified as potent, moderate or noninducers of CYP3A4 and CYP2B6 were tested. There was a good fit of data from upcyte\(^{®}\) hepatocytes to three different predictive models for CYP3A4 induction, namely the Relative Induction Score (RIS), AUC\(_{u}\)/F\(_{2}\), and C\(_{max,u}\)/Ind\(_{50}\). In addition, PXR (rifampicin) and CAR-selective (carbamazepine and phenytoin) inducers of CYP3A4 and CYP2B6 induction, respectively, were demonstrated. In conclusion, these data support the use of second-generation upcyte\(^{®}\) hepatocytes for CYP inhibition and induction assays. Under the culture conditions used, these cells expressed CYP activities that were equivalent to or higher than those measured in primary human hepatocyte cultures, which could be inhibited or induced by prototypical CYP inhibitors and inducers, respectively. Moreover, they can be used to predict in vivo CYP3A4 induction potential using three prediction models. Bulk availability of cells from multiple donors makes upcyte\(^{®}\) hepatocytes suitable for DDI screening, as well as more in-depth mechanistic investigations.
Converging evidence from controlled experiments suggests that the mere processing of a number and its attributes such as value or parity might affect free choice decisions between different actions. For example the spatial numerical associations of response codes (SNARC) effect indicates the magnitude of a digit to be associated with a spatial representation and might therefore affect spatial response choices (i.e., decisions between a "left" and a "right" option). At the same time, other (linguistic) features of a number such as parity are embedded into space and might likewise prime left or right responses through feature words [odd or even, respectively; markedness association of response codes (MARC) effect]. In this experiment we aimed at documenting such influences in a natural setting. We therefore assessed number space and parity space association effects by exposing participants to a fair distribution task in a card playing scenario. Participants drew cards, read out loud their number values, and announced their response choice, i.e., dealing it to a left vs. right player, indicated by Playmobil characters. Not only did participants prefer to deal more cards to the right player, the card's digits also affected response choices and led to a slightly but systematically unfair distribution, supported by a regular SNARC effect and counteracted by a reversed MARC effect. The experiment demonstrates the impact of SNARC- and MARC-like biases in free choice behavior through verbal and visual numerical information processing even in a setting with high external validity.
The main prediction of the Uncanny Valley Hypothesis (UVH) is that observation of humanlike characters that are difficult to distinguish from the human counterpart will evoke a state of negative affect. Well-established electrophysiological [late positive potential (LPP) and facial electromyography (EMG)] and self-report [Self-Assessment Manikin (SAM)] indices of valence and arousal, i.e., the primary orthogonal dimensions of affective experience, were used to test this prediction by examining affective experience in response to categorically ambiguous compared with unambiguous avatar and human faces (N = 30). LPP and EMG provided direct psychophysiological indices of affective state during passive observation and the SAM provided self-reported indices of affective state during explicit cognitive evaluation of static facial stimuli. The faces were drawn from well-controlled morph continua representing the UVH' dimension of human likeness (DHL). The results provide no support for the notion that category ambiguity along the DHL is specifically associated with enhanced experience of negative affect. On the contrary, the LPP and SAM-based measures of arousal and valence indicated a general increase in negative affective state (i.e., enhanced arousal and negative valence) with greater morph distance from the human end of the DHL. A second sample (N = 30) produced the same finding, using an ad hoc self-rating scale of feelings of familiarity, i.e., an oft-used measure of affective experience along the UVH' familiarity dimension. In conclusion, this multi-method approach using well-validated psychophysiological and self-rating indices of arousal and valence rejects for passive observation and for explicit affective evaluation of static faces the main prediction of the UVH.
Quantifying the spatio-temporal distribution of arthropods in tropical rainforests represents a first step towards scrutinizing the global distribution of biodiversity on Earth. To date most studies have focused on narrow taxonomic groups or lack a design that allows partitioning of the components of diversity. Here, we consider an exceptionally large dataset (113,952 individuals representing 5,858 species), obtained from the San Lorenzo forest in Panama, where the phylogenetic breadth of arthropod taxa was surveyed using 14 protocols targeting the soil, litter, understory, lower and upper canopy habitats, replicated across seasons in 2003 and 2004. This dataset is used to explore the relative influence of horizontal, vertical and seasonal drivers of arthropod distribution in this forest. We considered arthropod abundance, observed and estimated species richness, additive decomposition of species richness, multiplicative partitioning of species diversity, variation in species composition, species turnover and guild structure as components of diversity. At the scale of our study (2km of distance, 40m in height and 400 days), the effects related to the vertical and seasonal dimensions were most important. Most adult arthropods were collected from the soil/litter or the upper canopy and species richness was highest in the canopy. We compared the distribution of arthropods and trees within our study system. Effects related to the seasonal dimension were stronger for arthropods than for trees. We conclude that: (1) models of beta diversity developed for tropical trees are unlikely to be applicable to tropical arthropods; (2) it is imperative that estimates of global biodiversity derived from mass collecting of arthropods in tropical rainforests embrace the strong vertical and seasonal partitioning observed here; and (3) given the high species turnover observed between seasons, global climate change may have severe consequences for rainforest arthropods.
While interplay between BRCA1 and AURKA-RHAMM-TPX2-TUBG1 regulates mammary epithelial polarization, common genetic variation in HMMR (gene product RHAMM) may be associated with risk of breast cancer in BRCA1 mutation carriers. Following on these observations, we further assessed the link between the AURKA-HMMR-TPX2-TUBG1 functional module and risk of breast cancer in BRCA1 or BRCA2 mutation carriers. Forty-one single nucleotide polymorphisms (SNPs) were genotyped in 15,252 BRCA1 and 8,211 BRCA2 mutation carriers and subsequently analyzed using a retrospective likelihood approach. The association of HMMR rs299290 with breast cancer risk in BRCA1 mutation carriers was confirmed: per-allele hazard ratio (HR) = 1.10, 95% confidence interval (CI) 1.04 - 1.15, p = 1.9 x 10\(^{-4}\) (false discovery rate (FDR)-adjusted p = 0.043). Variation in CSTF1, located next to AURKA, was also found to be associated with breast cancer risk in BRCA2 mutation carriers: rs2426618 per-allele HR = 1.10, 95% CI 1.03 - 1.16, p = 0.005 (FDR-adjusted p = 0.045). Assessment of pairwise interactions provided suggestions (FDR-adjusted p\(_{interaction}\) values > 0.05) for deviations from the multiplicative model for rs299290 and CSTF1 rs6064391, and rs299290 and TUBG1 rs11649877 in both BRCA1 and BRCA2 mutation carriers. Following these suggestions, the expression of HMMR and AURKA or TUBG1 in sporadic breast tumors was found to potentially interact, influencing patients' survival. Together, the results of this study support the hypothesis of a causative link between altered function of AURKA-HMMR-TPX2-TUBG1 and breast carcinogenesis in BRCA1/2 mutation carriers.
Background
Autophagy participates in innate immunity by eliminating intracellular pathogens. Consequently, numerous microorganisms have developed strategies to impair the autophagic machinery in phagocytes. In the current study, interactions between Leishmania major (L. m.) and the autophagic machinery of bone marrow-derived macrophages (BMDM) were analyzed.
Methods
BMDM were generated from BALB/c mice, and the cells were infected with L. m. promastigotes. Transmission electron microscopy (TEM) and electron tomography were used to investigate the ultrastructure of BMDM and the intracellular parasites. Affymetrix® chip analyses were conducted to identify autophagy-related messenger RNAs (mRNAs) and microRNAs (miRNAs). The protein expression levels of autophagy related 5 (ATG5), BCL2/adenovirus E1B 19 kDa protein-interacting protein 3 (BNIP3), cathepsin E (CTSE), mechanistic target of rapamycin (MTOR), microtubule-associated proteins 1A/1B light chain 3B (LC3B), and ubiquitin (UB) were investigated through western blot analyses. BMDM were transfected with specific small interfering RNAs (siRNAs) against autophagy-related genes and with mimics or inhibitors of autophagy-associated miRNAs. The infection rates of BMDM were determined by light microscopy after a parasite-specific staining.
Results
The experiments demonstrated autophagy induction in BMDM after in vitro infection with L. m.. The results suggested a putative MTOR phosphorylation-dependent counteracting mechanism in the early infection phase and indicated that intracellular amastigotes were cleared by autophagy in BMDM in the late infection phase. Transcriptomic analyses and specific downregulation of protein expression with siRNAs suggested there is an association between the infection-specific over expression of BNIP3, as well as CTSE, and the autophagic activity of BMDM. Transfection with mimics of mmu-miR-101c and mmu-miR-129-5p, as well as with an inhibitor of mmu-miR-210-5p, demonstrated direct effects of the respective miRNAs on parasite clearance in L. m.-infected BMDM. Furthermore, Affymetrix® chip analyses revealed a complex autophagy-related RNA network consisting of differentially expressed mRNAs and miRNAs in BMDM, which indicates high glycolytic and inflammatory activity in the host macrophages.
Conclusions
Autophagy in L. m.-infected host macrophages is a highly regulated cellular process at both the RNA level and the protein level. Autophagy has the potential to clear parasites from the host. The results obtained from experiments with murine host macrophages could be translated in the future to develop innovative and therapeutic antileishmanial strategies for human patients.
Balanced hydroxyethylstarch (HES 130/0.4) impairs kidney function in-vivo without inflammation
(2015)
Volume therapy is a standard procedure in daily perioperative care, and there is an ongoing discussion about the benefits of colloid resuscitation with hydroxyethylstarch (HES). In sepsis HES should be avoided due to a higher risk for acute kidney injury (AKI). Results of the usage of HES in patients without sepsis are controversial. Therefore we conducted an animal study to evaluate the impact of 6% HES 130/0.4 on kidney integrity with sepsis or under healthy conditions Sepsis was induced by standardized Colon Ascendens Stent Peritonitis (sCASP). sCASP-group as well as control group (C) remained untreated for 24 h. After 18 h sCASP+HES group (sCASP+VOL) and control+HES (C+VOL) received 50 ml/KG balanced 6% HES (VOL) 130/0.4 over 6h. After 24h kidney function was measured via Inulin- and PAH-Clearance in re-anesthetized rats, and serum urea, creatinine (crea), cystatin C and Neutrophil gelatinase-associated lipocalin (NGAL) as well as histopathology were analysed. In vitro human proximal tubule cells (PTC) were cultured +/- lipopolysaccharid (LPS) and with 0.1–4.0% VOL. Cell viability was measured with XTT-, cell toxicity with LDH-test. sCASP induced severe septic AKI demonstrated divergent results regarding renal function by clearance or creatinine measure focusing on VOL. Soleley HES (C+VOL) deteriorated renal function without sCASP. Histopathology revealed significantly derangements in all HES groups compared to control. In vitro LPS did not worsen the HES induced reduction of cell viability in PTC cells. For the first time, we demonstrated, that application of 50 ml/KG 6% HES 130/0.4 over 6 hours induced AKI without inflammation in vivo. Severity of sCASP induced septic AKI might be no longer susceptible to the way of volume expansion
Behavioral profiles are influenced by both positive and negative experiences as well as the genetic disposition. Traditionally, accumulating adversity over lifetime is considered to predict increased anxiety like behavior ("allostatic load"). The alternative "mismatch hypothesis" suggests increased levels of anxiety if the early environment differs from the later-life environment. Thus, there is a need for a whole-life history approach to gain a deeper understanding of how behavioral profiles are shaped. The aim of this study was to elucidate the effects of life history on the behavioral profile of mice varying in serotonin transporter (5-HIT) genotype, an established mouse model of increased anxiety-like behavior. For this purpose, mice grew up under either adverse or beneficial conditions during early phases of life. In adulthood, they were further subdivided so as to face a situation that either matched or mismatched the condition experienced so far, resulting in four different life histories. Subsequently, mice were tested for their anxiety-like and exploratory behavior. The main results were: (1) Life history profoundly modulated the behavioral profile. Surprisingly, mice that experienced early beneficial and later escapable adverse conditions showed less anxiety-like and more exploratory behavior compared to mice of other life histories. (2) Genotype significantly influenced the behavioral profile, with homozygous 5-HTT knockout mice displaying highest levels of anxiety-like and lowest levels of exploratory behavior. Our findings concerning life history indicate that the absence of adversity does not necessarily cause lower levels of anxiety than accumulating adversity. Rather, some adversity may be beneficial, particularly when following positive events. Altogether, we conclude that for an understanding of behavioral profiles, it is not sufficient to look at experiences during single phases of life, but the whole life history has to be considered.
Marine sponge–associated actinomycetes are considered as promising sources for the discovery of novel biologically active compounds. In the present study, a total of 64 actinomycetes were isolated from 12 different marine sponge species that had been collected offshore the islands of Milos and Crete, Greece, eastern Mediterranean. The isolates were affiliated to 23 genera representing 8 different suborders based on nearly full length 16S rRNA gene sequencing. Four putatively novel species belonging to genera Geodermatophilus, Microlunatus, Rhodococcus and Actinomycetospora were identified based on a 16S rRNA gene sequence similarity of < 98.5% to currently described strains. Eight actinomycete isolates showed bioactivities against Trypanosma brucei brucei TC221 with half maximal inhibitory concentration (IC50) values <20 μg/mL. Thirty four isolates from the Milos collection and 12 isolates from the Crete collection were subjected to metabolomic analysis using high resolution LC-MS and NMR for dereplication purposes. Two isolates belonging to the genera Streptomyces (SBT348) and Micromonospora (SBT687) were prioritized based on their distinct chemistry profiles as well as their anti-trypanosomal activities. These findings demonstrated the feasibility and efficacy of utilizing metabolomics tools to prioritize chemically unique strains from microorganism collections and further highlight sponges as rich source for novel and bioactive actinomycetes.
In this study, the ability of a multiwalled carbon nanotube functionalized with fluorescein isothiocyanate (MWCNT-FITC) was assessed as a prospective central nervous system-targeting drug delivery system to permeate the blood-brain barrier. The results indicated that the MWCNT-FITC conjugate is able to penetrate microvascular cerebral endothelial monolayers; its concentrations in the Transwell® system were fully equilibrated after 48 hours. Cell viability test, together with phase-contrast and fluorescence microscopies, did not detect any signs of MWCNT-FITC toxicity on the cerebral endothelial cells. These microscopic techniques also revealed presumably the intracellular localization of fluorescent MWCNT-FITCs apart from their massive nonfluorescent accumulation on the cellular surface due to nanotube lipophilic properties. In addition, the 1,000 ps molecular dynamics simulation in vacuo discovered the phenomenon of carbon nanotube aggregation driven by van der Waals forces via MWCN-TFITC rapid dissociation as an intermediate phase.
The novel BackHome system offers individuals with disabilities a range of useful services available via brain-computer interfaces (BCIs), to help restore their independence. This is the time such technology is ready to be deployed in the real world, that is, at the target end users’ home. This has been achieved by the development of practical electrodes, easy to use software, and delivering telemonitoring and home support capabilities which have been conceived, implemented, and tested within a user-centred design approach. The final BackHome system is the result of a 3-year long process involving extensive user engagement to maximize effectiveness, reliability, robustness, and ease of use of a home based BCI system. The system is comprised of ergonomic and hassle-free BCI equipment; one-click software services for Smart Home control, cognitive stimulation, and web browsing; and remote telemonitoring and home support tools to enable independent home use for nonexpert caregivers and users. BackHome aims to successfully bring BCIs to the home of people with limited mobility to restore their independence and ultimately improve their quality of life.
The active zone (AZ) protein Bruchpilot (Brp) is essential for rapid glutamate release at Drosophila melanogaster neuromuscular junctions (NMJs). Quantal time course and measurements of action potential-waveform suggest that presynaptic fusion mechanisms are altered in brp null mutants (brp\(^{69}\)). This could account for their increased evoked excitatory postsynaptic current (EPSC) delay and rise time (by about 1 ms). To test the mechanism of release protraction at brp\(^{69}\) AZs, we performed knock-down of Synaptotagmin-1 (Syt) via RNAi (syt\(^{KD}\)) in wildtype (wt), brp\(^{69}\) and rab3 null mutants (rab3\(^{rup}\)), where Brp is concentrated at a small number of AZs. At wt and rab3\(^{rup}\) synapses, syt\(^{KD}\) lowered EPSC amplitude while increasing rise time and delay, consistent with the role of Syt as a release sensor. In contrast, syt\(^{KD}\) did not alter EPSC amplitude at brp\(^{69}\) synapses, but shortened delay and rise time. In fact, following syt\(^{KD}\), these kinetic properties were strikingly similar in wt and brp\(^{69}\), which supports the notion that Syt protracts release at brp\(^{69}\) synapses. To gain insight into this surprising role of Syt at brp\(^{69}\) AZs, we analyzed the structural and functional differentiation of synaptic boutons at the NMJ. At tonic type Ib motor neurons, distal boutons contain more AZs, more Brp proteins per AZ and show elevated and accelerated glutamate release compared to proximal boutons. The functional differentiation between proximal and distal boutons is Brp-dependent and reduced after syt\(^{KD}\). Notably, syt\(^{KD}\) boutons are smaller, contain fewer Brp positive AZs and these are of similar number in proximal and distal boutons. In addition, super-resolution imaging via dSTORM revealed that syt\(^{KD}\) increases the number and alters the spatial distribution of Brp molecules at AZs, while the gradient of Brp proteins per AZ is diminished. In summary, these data demonstrate that normal structural and functional differentiation of Drosophila AZs requires concerted action of Brp and Syt.
Background
Suture pretension during tendon repair is supposed to increase the resistance to gap formation. However, its effects on the Bunnell suture technique are unknown. The purpose of this study was to determine the biomechanical effects of suture pretension on the Bunnell and cross-lock Bunnell techniques for tendon repair.
Methods
Eighty porcine hindlimb tendons were randomly assigned to four different tendon repair groups: those repaired with or without suture pretension using either a simple Bunnell or cross-lock Bunnell technique. Pretension was applied as a 10 % shortening of the sutured tendon. After measuring the cross-sectional diameter at the repair site, static and cyclic biomechanical tests were conducted to evaluate the initial and 5-mm gap formation forces, elongation during cyclic loading, maximum tensile strength, and mode of failure. The suture failure mechanism was also separately assessed fluoroscopically in two tendons that were repaired with steel wire.
Results
Suture pretension was accompanied by a 10 to 15 % increase in the tendon diameter at the repair site. Therefore, suture pretension with the Bunnell and cross-lock Bunnell repair techniques noticeably increased the resistance to initial gap formation and 5-mm gap formation. The tension-free cross-lock Bunnell repair demonstrated more resistance to initial and 5-mm gap formation, less elongation, and higher maximum tensile strength than the tension-free Bunnell repair technique. The only difference between the tensioned cross-lock Bunnell and tensioned Bunnell techniques was a larger resistance to 5-mm gap formation with the cross-lock Bunnell technique. Use of the simple instead of cross-lock suture configuration led to failure by suture cut out, as demonstrated fluoroscopically.
Conclusion
Based on these results, suture pretension decreases gapping and elongation after tendon repair, and those effects are stronger when using a cross-lock, rather than a regular Bunnell suture. However, pretension causes an unfavorable increase in the tendon diameter at the repair site, which may adversely affect wound healing.
Cadherin-13 (CDH13), a unique glycosylphosphatidylinositol-anchored member of the cadherin family of cell adhesion molecules, has been identified as a risk gene for attention-deficit/hyperactivity disorder (ADHD) and various comorbid neurodevelopmental and psychiatric conditions, including depression, substance abuse, autism spectrum disorder and violent behavior, while the mechanism whereby CDH13 dysfunction influences pathogenesis of neuropsychiatric disorders remains elusive. Here we explored the potential role of CDH13 in the inhibitory modulation of brain activity by investigating synaptic function of GABAergic interneurons. Cellular and subcellular distribution of CDH13 was analyzed in the murine hippocampus and a mouse model with a targeted inactivation of Cdh13 was generated to evaluate how CDH13 modulates synaptic activity of hippocampal interneurons and behavioral domains related to psychopathologic (endo) phenotypes. We show that CDH13 expression in the cornu ammonis (CA) region of the hippocampus is confined to distinct classes of interneurons. Specifically, CDH13 is expressed by numerous parvalbumin and somatostatin-expressing interneurons located in the stratum oriens, where it localizes to both the soma and the presynaptic compartment. Cdh13\(^{-/-}\) mice show an increase in basal inhibitory, but not excitatory, synaptic transmission in CA1 pyramidal neurons. Associated with these alterations in hippocampal function, Cdh13\(^{-/-}\) mice display deficits in learning and memory. Taken together, our results indicate that CDH13 is a negative regulator of inhibitory synapses in the hippocampus, and provide insights into how CDH13 dysfunction may contribute to the excitatory/inhibitory imbalance observed in neurodevelopmental disorders, such as ADHD and autism.
Juvenile Dermatomyositis (JDM) is a rare autoimmune disease in children and adolescents. In these patients calcinosis might be the most characteristic symptom. However there are only few reported cases of intramuscular calcinosis in Dermatomyositis. We report a case of calcinosis universalis (CU) of the elbow in JDM successfully treated with broaching. The patient, a 24-year-old woman, suffered from a long history of JDM. On examination she presented with a fistula lateral to the olecranon and pain of the right elbow joint. Plain X-rays displayed a diffuse pattern of multiple periarticular, subcutaneous, and intramuscular calcifications. The patient underwent surgery for histological and microbiological sampling as well as broaching. Intraoperatively sinus formation and subfascial hard calcium deposition were found. Due to the risk of collateral tissue damage, incomplete broaching was performed. A local infection with Staphylococcus was diagnosed and treated with antibiotics. On six-week and 30-month follow-up the patient was free of pain and had very good function. Calcifications on standard radiographs had almost resolved entirely. This case report gives a summary on calcinosis in Dermatomyositis and adds a new case of recalcitrant CU to the literature. Broaching surgery proved to be a reliable treatment option in symptomatic calcinosis.
DNA double strand break (DSB) formation induced by ionizing radiation exposure is indicated by the DSB biomarkers \(\gamma\)-H2AX and 53BP1. Knowledge about DSB foci formation in-vitro after internal irradiation of whole blood samples with radionuclides in solution will help us to gain detailed insights about dose-response relationships in patients after molecular radiotherapy (MRT). Therefore, we studied the induction of radiation-induced co-localizing \(\gamma\)-H2AX and 53BP1 foci as surrogate markers for DSBs in-vitro, and correlated the obtained foci per cell values with the in-vitro absorbed doses to the blood for the two most frequently used radionuclides in MRT (I-131 and Lu-177). This approach led to an in-vitro calibration curve. Overall, 55 blood samples of three healthy volunteers were analyzed. For each experiment several vials containing a mixture of whole blood and radioactive solutions with different concentrations of isotonic NaCl-diluted radionuclides with known activities were prepared. Leukocytes were recovered by density centrifugation after incubation and constant blending for 1 h at 37°C. After ethanol fixation they were subjected to two-color immunofluorescence staining and the average frequencies of the co-localizing \(\gamma\)-H2AX and 53BP1 foci/nucleus were determined using a fluorescence microscope equipped with a red/green double band pass filter. The exact activity was determined in parallel in each blood sample by calibrated germanium detector measurements. The absorbed dose rates to the blood per nuclear disintegrations occurring in 1 ml of blood were calculated for both isotopes by a Monte Carlo simulation. The measured blood doses in our samples ranged from 6 to 95 mGy. A linear relationship was found between the number of DSB-marking foci/nucleus and the absorbed dose to the blood for both radionuclides studied. There were only minor nuclide-specific intra-and inter-subject deviations.
Background
Pneumonia frequently complicates stroke and has amajor impact on outcome. We derived and internally validated a simple clinical risk score for predicting stroke-associated pneumonia (SAP), and compared the performance with an existing score (A\(^{2}\)DS\(^{2}\)).
Methods and Results
We extracted data for patients with ischemic stroke or intracerebral hemorrhage from the Sentinel Stroke National Audit Programme multicenter UK registry. The data were randomly allocated into derivation (n=11 551) and validation (n=11 648) samples. A multivariable logistic regression model was fitted to the derivation data to predict SAP in the first 7 days of admission. The characteristics of the score were evaluated using receiver operating characteristics (discrimination) and by plotting predicted versus observed SAP frequency in deciles of risk (calibration). Prevalence of SAP was 6.7% overall. The final 22-point score (ISAN: prestroke Independence [modified Rankin scale], Sex, Age, National Institutes of Health Stroke Scale) exhibited good discrimination in the ischemic stroke derivation (C-statistic 0.79; 95% CI 0.77 to 0.81) and validation (C-statistic 0.78; 95% CI 0.76 to 0.80) samples. It was well calibrated in ischemic stroke and was further classified into meaningful risk groups (low 0 to 5, medium6 to 10, high 11 to 14, and very high >= 15) associated with SAP frequencies of 1.6%, 4.9%, 12.6%, and 26.4%, respectively, in the validation sample. Discrimination for both scores was similar, although they performed less well in the intracerebral hemorrhage patients with an apparent ceiling effect.
Conclusions
The ISAN score is a simple tool for predicting SAP in clinical practice. External validation is required in ischemic and hemorrhagic stroke cohorts.
Stimulating the immune system to attack cancer is a promising approach, even for the control of advanced cancers. Several cytokines that promote interferon-γ-dominated immune responses show antitumor activity, with interleukin 12 (IL-12) being of major importance. Here, we used an antibody-IL-12 fusion protein (NHS-IL12) that binds histones of necrotic cells to treat human sarcoma in humanized mice. Following sarcoma engraftment, NHS-IL12 therapy was combined with either engineered IL-7 (FcIL-7) or IL-2 (IL-2MAB602) for continuous cytokine bioavailability. NHS-IL12 strongly induced innate and adaptive antitumor immunity when combined with IL-7 or IL-2. NHS-IL12 therapy significantly improved survival of sarcoma-bearing mice and caused long-term remissions when combined with IL-2. NHS-IL12 induced pronounced cancer cell senescence, as documented by strong expression of senescence-associated p16\(^{INK4a}\) and nuclear translocation of p-HP1γ, and permanent arrest of cancer cell proliferation. In addition, this cancer immunotherapy initiated the induction of myogenic differentiation, further promoting the hypothesis that efficient antitumor immunity includes mechanisms different from cytotoxicity for efficient cancer control in vivo.
Background
Animal models have been instrumental in defining thrombus formation, including the role of platelet surface glycoprotein (GP) receptors, in acute ischemic stroke (AIS). However, the involvement of GP receptors in human ischemic stroke pathophysiology and their utility as biomarkers for ischemic stroke risk and severity requires elucidation.
Aims
To determine whether platelet GPIb and GPIIb/IIIa receptors are differentially expressed in patients with AIS and chronic cerebrovascular disease (CCD) compared with healthy volunteers (HV) and to identify predictors of GPIb and GPIIb/IIIa expression.
Methods
This was a case-control study of 116 patients with AIS or transient ischemic attack (TIA), 117 patients with CCD, and 104 HV who were enrolled at our University hospital from 2010 to 2013. Blood sampling was performed once in the CCD and HV groups, and at several time points in patients with AIS or TIA. Linear regression and analysis of variance were used to analyze correlations between platelet GPIb and GPIIb/IIIa receptor numbers and demographic and clinical parameters.
Results
GPIb and GPIIb/IIIa receptor numbers did not significantly differ between the AIS, CCD, and HV groups. GPIb receptor expression level correlated significantly with the magnitude of GPIIb/IIIa receptor expression and the neutrophil count. In contrast, GPIIb/IIIa receptor numbers were not associated with peripheral immune-cell sub-population counts. Creactive protein was an independent predictor of GPIIb/IIIa (not GPIb) receptor numbers.
Conclusions
Platelet GPIb and GPIIb/IIIa receptor numbers did not distinguish between patient or control groups in this study, negating their potential use as a biomarker for predicting stroke risk.
Influenza A virus (IAV) infection causes an acute respiratory disease characterized by a strong inflammatory immune response and severe immunopathology. Proinflammatory mechanisms are well described in the murine IAV infection model, but less is known about the mechanisms leading to the resolution of inflammation. Here, we analyzed the contribution of CD11b\(^{+}\)Ly6C\(^{++}\)Ly6G\(^{-}\) cells to this process. An accumulation of CD11b\(^{+}\)Ly6C\(^{++}\)Ly6G\(^{-}\) cells within the lungs was observed during the course of IAV infection. Phenotypic characterization of these CD11b\(^{+}\)Ly6C\(^{++}\)Ly6G\(^{-}\) cells by flow cytometry and RNA-Seq revealed an activated phenotype showing both pro- and anti-inflammatory features, including the expression of inducible nitric oxide synthase (iNOS) by a fraction of cells in an IFN-γ-dependent manner. Moreover, CD11b\(^{+}\)Ly6C\(^{++}\)Ly6G\(^{-}\) cells isolated from lungs of IAV-infected animals displayed suppressive activity when tested in vitro, and iNOS inhibitors could abrogate this suppressive activity. Collectively, our data suggest that during IAV infection, CD11b\(^{+}\)Ly6C\(^{++}\)Ly6G\(^{-}\) cells acquire immunoregulatory function, which might contribute to the prevention of pathology during this life-threatening disease.
Introduction
Enhanced B cell activity, particularly memory B cells have gained interest in evaluating response during therapies with biologics. CD27-IgD- double-negative (DN) B cells lacking the conventional memory marker CD27 are reported to be part of the memory compartment, however, only scarce data is available for rheumatoid arthritis (RA). We therefore focused on DN B cells in RA, studied their isotypes and modulation during interleukin-6 receptor (IL-6R) inhibition by tocilizumab (TCZ).
Methods
DN B cells were phenotypically analyzed from 40 RA patients during TCZ at baseline week 12, week 24 and 1 year. A single B cell polymerase chain reaction (PCR) approach was used to study Ig receptors, VH gene rearrangements and specific isotypes.
Results
Phenotypic analysis showed a significantly expanded population of DN B cells in RA which contain a heterogeneous mixture of IgG-, IgA- and IgM-expressing cells with a clear dominance of IgG+ cells. DN B cells carry rearranged heavy chain gene sequences with a diversified mutational pattern consistent with memory B cells. In contrast to tumor necrosis factor alpha (TNF-α) inhibition, a significant reduction in mutational frequency of BCR gene rearrangements at week 12, 24 and 1 year (P <0.0001) was observed by in vivo IL-6R inhibition. These changes were observed for all BCR isotypes IgG, IgA and IgM at week 12, 24 and 1 year (P <0.0001). IgA-RF, IgA serum level and IgA+ DN B cells decreased significantly (P <0.05) at week 12 and week 24 during TCZ. Patients with a good European League Against Rheumatism (EULAR) response to TCZ had less DN B cells at baseline as compared to moderate responders (P = 0.006). Univariate logistic regression analysis revealed that the frequency of DN B cells at baseline is inversely correlated to a subsequent good EULAR response (P = 0.024) with an odds ratio of 1.48 (95% confidence interval as 1.05 to 2.06).
Conclusions
In RA, the heterogeneous DN B cell compartment is expanded and dominated by IgG isotype. TCZ can modulate the mutational status of DN Ig isotype receptors over 1 year. Interestingly, the frequency of DN B cells in RA may serve as a baseline predictor of subsequent EULAR response to TCZ.
Measurements of the centrality and rapidity dependence of inclusive jet production in \(\sqrt{^SNN}\)=5.02 TeV proton–lead (p+Pb) collisions and the jet cross-section in \(\sqrt{s}\)=2.76 TeV proton–proton collisions are presented. These quantities are measured in datasets corresponding to an integrated luminosity of 27.8 nb\(^{−1}\) and 4.0 pb\(^{−1}\), respectively, recorded with the ATLAS detector at the Large Hadron Collider in 2013. The p+Pb collision centrality was characterised using the total transverse energy measured in the pseudorapidity interval −4.9<η<−3.2 in the direction of the lead beam. Results are presented for the double-differential per-collision yields as a function of jet rapidity and transverse momentum (\(p_T\)) for minimum-bias and centrality-selected p+Pb collisions, and are compared to the jet rate from the geometric expectation. The total jet yield in minimum-bias events is slightly enhanced above the expectation in a \(p_T\)-dependent manner but is consistent with the expectation within uncertainties. The ratios of jet spectra from different centrality selections show a strong modification of jet production at all \(p_T\) at forward rapidities and for large \(p_T\) at mid-rapidity, which manifests as a suppression of the jet yield in central events and an enhancement in peripheral events. These effects imply that the factorisation between hard and soft processes is violated at an unexpected level in proton–nucleus collisions. Furthermore, the modifications at forward rapidities are found to be a function of the total jet energy only, implying that the violations may have a simple dependence on the hard parton–parton kinematics.
Background
The aim is to analyze characteristics and to study the potentials of non-coplanar intensity modulated radiation therapy (IMRT) techniques. The planning study applies to generalized organ at risk (OAR) – planning target volume (PTV) geometries.
Methods
The authors focus on OARs embedded in the PTV. The OAR shapes are spherically symmetric (A), cylindrical (B), and bended (C). Several IMRT techniques are used for the planning study: a) non-coplanar quasi-isotropic; b) two sets of equidistant coplanar beams, half of beams incident in a plane perpendicular to the principal plane; c) coplanar equidistant (reference); d) coplanar plus one orthogonal beam. The number of beam directions varies from 9 to 16. The orientation of the beam sets is systematically changed; dose distributions resulting from optimal fluence are explored. A selection of plans is optimized with direct machine parameter optimization (DMPO) allowing 120 and 64 segments. The overall plan quality, PTV coverage, and OAR sparing are evaluated.
Results
For all fluence based techniques in cases A and C, plan quality increased considerably if more irradiation directions were used. For the cylindrically symmetric case B, however, only a weak beam number dependence was observed for the best beam set orientation, for which non-coplanar directions could be found where OAR- and PTV-projections did not overlap. IMRT plans using quasi-isotropical distributed non-coplanar beams showed stable results for all topologies A, B, C, as long as 16 beams were chosen; also the most unfavorable beam arrangement created results of similar quality as the optimally oriented coplanar configuration. For smaller number of beams or application in the trunk, a coplanar technique with additional orthogonal beam could be recommended. Techniques using 120 segments created by DMPO could qualitatively reproduce the fluence based results. However, for a reduced number of segments the beam number dependence declined or even reversed for the used planning system and the plan quality degraded substantially.
Conclusions
Topologies with targets encompassing sensitive OAR require sufficient number of beams of 15 or more. For the subgroup of topologies where beam incidences are possible which cover the whole PTV without direct OAR irradiation, the quality dependence on the number of beams is much less pronounced above 9 beams. However, these special non-coplanar beam directions have to be found. On the basis of this work the non-coplanar IMRT techniques can be chosen for further clinical planning studies.
Introduction
The bursa subacromialis (BS) provides the gliding mechanism of the shoulder and regenerates itself after surgical removal. Therefore, we explored the presence of mesenchymal stem cells (MSCs) within the human adult BS tissue and characterized the BS cells compared to MSCs from bone marrow (BMSCs) on a molecular level.
Methods
BS cells were isolated by collagenase digest from BS tissues derived from patients with degenerative rotator cuff tears, and BMSCs were recovered by adherent culture from bone-marrow of patients with osteoarthritis of the hip. BS cells and BMSCs were compared upon their potential to proliferate and differentiate along chondrogenic, osteogenic and adipogenic lineages under specific culture conditions. Expression profiles of markers associated with mesenchymal phenotypes were comparatively evaluated by flow cytometry, immunohistochemistry, and whole genome array analyses.
Results
BS cells and BMSCs appeared mainly fibroblastic and revealed almost similar surface antigen expression profiles, which was \(CD44^+, CD73^+, CD90^+, CD105^+, CD106^+\),\(STRO-1^+, CD14^−, CD31^−, CD34^− ,
CD45^−, CD144^−\). Array analyses revealed 1969 genes upregulated and 1184 genes downregulated in BS cells vs. BMSCs, indicating a high level of transcriptome similarity. After 3 weeks of differentiation culture, BS cells and BMSCs showed a similar strong chondrogenic, adipogenic and osteogenic potential, as shown by histological, immunohistochemical and RT-PCR analyses in contrast to the respective negative controls.
Conclusions
Our in vitro characterizations show that BS cells fulfill all characteristics of mesenchymal stem cells, and therefore merit further attention for the development of improved therapies for various shoulder pathologies.
Background
The spontaneously hypertensive rat (SHR) has been used to model changes in the central nervous system associated with cognitive-related disorders. Recent human and animal studies indicate a possible relationship between cognitive deficits, insulin resistance and hypertension. We aimed to investigate whether cognitively impaired SHRs develop central and/or peripheral insulin resistance and how their cognitive performance is influenced by the animal’s sex and age as well as strains used for comparison (Wistar and Wistar-Kyoto/WKY).
Methods
Three and seven-month-old SHR, Wistar, and WKY rats were studied for their cognitive performance using Morris Water Maze (MWM) and Passive Avoidance tests (PAT). Plasma glucose and insulin were obtained after oral glucose tolerance tests. Cerebral cortex, hippocampus, and striatum status of insulin-receptor (IR) β-subunit and glycogen synthase kinase-3β (GSK3β) and their phosphorylated forms were obtained via ELISA.
Results
SHRs performed poorly in MWM and PAT in comparison to both control strains but more pronouncedly compared to WKY. Females performed poorer than males and 7-month-old SHRs had poorer MWM performance than 3-month-old ones. Although plasma glucose levels remained unchanged, plasma insulin levels were significantly increased in the glucose tolerance test in 7-month-old SHRs. SHRs demonstrated reduced expression and increased activity of IRβ-subunit in cerebral cortex, hippocampus, and striatum with different regional changes in phospho/total GSK3β ratio, as compared to WKYs.
Conclusion
Results indicate that cognitive deficits in SHRs are accompanied by both central and peripheral insulin dysfunction, thus allowing for the speculation that SHRs might additionally be considered as a model of insulin resistance-induced type of dementia.
Immune cells (IC) play a crucial role in murine stroke pathophysiology. However, data are limited on the role of these cells in ischemic stroke in humans. We therefore aimed to characterize and compare peripheral IC subsets in patients with acute ischemic stroke/transient ischemic attack (AIS/TIA), chronic cerebrovascular disease (CCD) and healthy volunteers (HV). We conducted a case-control study of patients with AIS/TIA (n = 116) or CCD (n = 117), and HV (n = 104) who were enrolled at the University Hospital Würzburg from 2010 to 2013. We determined the expression and quantity of IC subsets in the three study groups and performed correlation analyses with demographic and clinical parameters. The quantity of several IC subsets differed between the AIS/TIA, CCD, and HV groups. Several clinical and demographic variables independently predicted the quantity of IC subsets in patients with AIS/TIA. No significant changes in the quantity of IC subsets occurred within the first three days after AIS/TIA. Overall, these findings strengthen the evidence for a pathophysiologic role of IC in human ischemic stroke and the potential use of IC-based biomarkers for the prediction of stroke risk. A comprehensive description of IC kinetics is crucial to enable the design of targeted treatment strategies.
Most humans become infected with human cytomegalovirus (HCMV). Typically, the immune system controls the infection, but the virus persists and can reactivate in states of immunodeficiency. While substantial information is available on the contribution of CD8 T cells and antibodies to anti-HCMV immunity, studies of the T\(_{H}\)1, T\(_{H}\)2, and T\(_{H}\)17 subsets have been limited by the low frequency of HCMV-specific CD4 T cells in peripheral blood mononuclear cell (PBMC). Using the enzyme-linked Immunospot\(^{®}\) assay (ELISPOT) that excels in low frequency measurements, we have established these in a sizable cohort of healthy HCMV controllers. Cytokine recall responses were seen in all seropositive donors. Specifically, interferon (IFN)-\({\gamma}\) and/or interleukin (IL)-17 were seen in isolation or with IL-4 in all test subjects. IL-4 recall did not occur in isolation. While the ratios of T\(_{H}\)1, T\(_{H}\)2, and T\(_{H}\)17 cells exhibited substantial variations between different individuals these ratios and the frequencies were relatively stable when tested in samples drawn up to five years apart. IFN-\({\gamma}\) and IL-2 co-expressing polyfunctional cells were seen in most subjects. Around half of the HCMV-specific CD4 cells were in a reversible state of exhaustion. The data provided here established the T\(_{H}\)1, T\(_{H}\)2, and T\(_{H}\)17 characteristic of the CD4 cells that convey immune protection for successful immune surveillance against which reactivity can be compared when the immune surveillance of HCMV fails.
The mouse gastro-intestinal and biliary tract mucosal epithelia harbor choline acetyltransferase (ChAT)-positive brush cells with taste cell-like traits. With the aid of two transgenic mouse lines that express green fluorescent protein (EGFP) under the control of the ChAT promoter (EGFP\(^{ChAT}\)) and by using in situ hybridization and immunohistochemistry we found that EGFP\(^{ChAT}\) cells were clustered in the epithelium lining the gastric groove. EGFP\(^{ChAT}\) cells were numerous in the gall bladder and bile duct, and found scattered as solitary cells along the small and large intestine. While all EGFP\(^{ChAT}\) cells were also ChAT-positive, expression of the high-affinity choline transporter (ChT1) was never detected. Except for the proximal colon, EGFP\(^{ChAT}\) cells also lacked detectable expression of the vesicular acetylcholine transporter (VAChT). EGFP\(^{ChAT}\) cells were found to be separate from enteroendocrine cells, however they were all immunoreactive for cytokeratin 18 (CK18), transient receptor potential melastatin-like subtype 5 channel (TRPM5), and for cyclooxygenases 1 (COX1) and 2 (COX2). The ex vivo stimulation of colonic EGFP\(^{ChAT}\) cells with the bitter substance denatonium resulted in a strong increase in intracellular calcium, while in other epithelial cells such an increase was significantly weaker and also timely delayed. Subsequent stimulation with cycloheximide was ineffective in both cell populations. Given their chemical coding and chemosensory properties, EGFP\(^{ChAT}\) brush cells thus may have integrative functions and participate in induction of protective reflexes and inflammatory events by utilizing ACh and prostaglandins for paracrine signaling.
Mitotic chromosomes of 16 species of the frog genus Xenopus were prepared from kidney and lung cell cultures. In the chromosomes of 7 species, high-resolution replication banding patterns could be induced by treating the cultures with 5-bromodeoxyuridine (BrdU) and deoxythymidine (dT) in succession, and in 6 of these species the BrdU/dT-banded chromosomes could be arranged into karyotypes. In the 3 species of the clade with 2n = 20 and 4n = 40 chromosomes (X. tropicalis, X. epitropicalis, X. new tetraploid 1), as well as in the 3 species with 4n = 36 chromosomes (X. laevis, X. borealis, X. muelleri), the BrdU/dT-banded karyotypes show a high degree of homoeology, though differences were detected between these groups. Translocations, inversions, insertions or sex-specific replication bands were not observed. Minor replication asynchronies found between chromosomes probably involve heterochromatic regions. BrdU/dT replication banding of Xenopus chromosomes provides the landmarks necessary for the exact physical mapping of genes and repetitive sequences. FISH with an X. laevis 5S rDNA probe detected multiple hybridization sites at or near the long-arm telomeric regions in most chromosomes of X. laevis and X. borealis, whereas in X. muelleri, the 5S rDNA sequences are located exclusively at the long-arm telomeres of a single chromosome pair. Staining with the AT base pair-specific fluorochrome quinacrine mustard revealed brightly fluorescing heterochromatic regions in the majority of X. borealis chromosomes which are absent in other Xenopus species.
Background
Chronic hyperglycemia (CHG) with HbA1c as an indicator affects postoperative mortality and morbidity after coronary artery bypass grafting surgery (CABG). Acute kidney injury (AKI) is one of the frequent postoperative complications after CABG impacting short-and long-term outcomes. We investigated the association between CHG and postoperative incidence of AKI in CABG patients with and without history of diabetes mellitus (DM).
Methods
This cohort study consecutively enrolled patients undergoing CABG in 2009 at the department for cardiovascular surgery. CHG was defined as HbA1c ≥ 6.0 %. Patients with advanced chronic kidney disease (CKD) were excluded. The incidence of postoperative AKI and its association with CHG was analyzed by univariate and multivariate logistic regression modeling.
Results
Three-hundred-seven patients were analyzed. The incidence of AKI was 48.2 %. Patients with CHG (n = 165) were more likely to be female and had greater waist circumference as well as other comorbid conditions, such as smoking, history of DM, CKD, hypertension, pulmonary hypertension, and chronic obstructive pulmonary disease (all p ≤ 0.05). Preoperative eGFR, atrial fibrillation (AF), history of DM and CHG were associated with an increased risk of postoperative AKI in univariate analyses. In multivariate modelling, history of DM as well as preoperative eGFR and AF lost significance, while age, CHG and prolonged OP duration (p < 0.05) were independently associated with postoperative AKI.
Conclusions
Our results suggest that CHG defined on a single measurement of HbA1c ≥ 6.0 % was associated with the incidence of AKI after CABG. This finding might implicate that treatment decisions, including the selection of operative strategies, could be based on HbA1c measurement rather than on a recorded history of diabetes.
Human leishmaniasis covers a broad spectrum of clinical manifestations ranging from self-healing cutaneous leishmaniasis to severe and lethal visceral leishmaniasis caused among other species by Leishmania major or Leishmania donovani, respectively. Some drug candidates are in clinical trials to substitute current therapies, which are facing emerging drug-resistance accompanied with serious side effects. Here, two cinnamic acid bornyl ester derivatives (1 and 2) were assessed for their antileishmanial activity. Good selectivity and antileishmanial activity of bornyl 3-phenylpropanoate (2) in vitro prompted the antileishmanial assessment in vivo. For this purpose, BALB/c mice were infected with Leishmania major promastigotes and treated with three doses of 50 mg/kg/day of compound 2. The treatment prevented the characteristic swelling at the site of infection and correlated with reduced parasite burden. Transmitted light microscopy and transmission electron microscopy of Leishmania major promastigotes revealed that compounds 1 and 2 induce mitochondrial swelling. Subsequent studies on Leishmania major promastigotes showed the loss of mitochondrial transmembrane potential (ΔΨm) as a putative mode of action. As the cinnamic acid bornyl ester derivatives 1 and 2 had exhibited antileishmanial activity in vitro, and compound 2 in Leishmania major-infected BALB/c mice in vivo, they can be regarded as possible lead structures for the development of new antileishmanial therapeutic approaches.
Incremental exercise testing is frequently used as a tool for evaluating determinants of endurance performance. The available reference values for the peak oxygen uptake \((VO_{2peak})\), % of \(VO_{2peak}\) , running speed at the lactate threshold \((v_{LT})\), running economy (RE), and maximal running speed \((v_{peak})\) for different age, gender, and disciplines are not sufficient for the elite athletic population. The key variables of 491 young athletes (age range 12–21 years; 250 males, 241 females) assessed during a running step test protocol \((2.4 m s^{−1} ; increase 0.4 m s^{−1} 5 min^{−1})\) were analysed in five subgroups, which were related to combat-, team-, endurance-, sprint- and power-, and racquet-related disciplines. Compared with female athletes, male athletes achieved a higher \(v_{peak}\) (P = 0.004). The body mass, lean body mass, height, abs. \(VO_{2peak} (ml min^{−1})\), rel. \(VO_{2peak} (ml kg^{−1} min^{−1})\), rel. \(VO_{2peak} (ml min^{−1} kg^{−0.75})\), and RE were higher in the male participants compared with the females (P < 0.01). The % of \(VO_2\) at \(v_{LT}\) was lower in the males compared with the females (P < 0.01). No differences between gender were detected for the \(v_{LT}\) (P = 0.17) and % of \(VO_2\) at \(v_{LT}\) (P = 0.42). This study is one of the first to provide a broad spectrum of data to classify nearly 500 elite athletes aged 12–21 years of both gender and different disciplines.
Background:
We assessed the diagnostic value of standard clinical methods and combined biomarker testing (galactomannan assay and polymerase chain reaction screening) in a prospective case-control study to detect invasive pulmonary aspergillosis in patients with hematological malignancies and prolonged neutropenia.
Methods:
In this observational study 162 biomarker analyses were performed on samples from 27 febrile neutropenic episodes. Sera were successively screened for galactomannan antigen and for Aspergillus fumigatus specific nucleic acid targets. Furthermore thoracic computed tomography scanning was performed along with bronchoscopy with lavage when clinically indicated. Patients were retrospectively stratified to define a case-group with "proven" or "probable" invasive pulmonary aspergillosis (25.93 %) and a control-group of patients with no evidence for of invasive pulmonary aspergillosis (74.07 %). In 44.44 % of episodes fever ceased in response to antibiotic treatment (group II). Empirical antifungal therapy was administered for episodes with persistent or relapsing fever (group I). 48.15 % of patients died during the study period. Postmortem histology was pursued in 53.85 % of fatalities.
Results:
Concordant negative galactomannan and computed tomography supported by a polymerase chain reaction assay were shown to have the highest discriminatory power to exclude invasive pulmonary aspergillosis. Bronchoalveolar lavage was performed in 6 cases of invasive pulmonary aspergillosis and in 15 controls. Although bronchoalveolar lavage proved negative in 93 % of controls it did not detect IPA in 86 % of the cases. Remarkably post mortem histology convincingly supported the presence of Aspergillus hyphae in lung tissue from a single case which had consecutive positive polymerase chain reaction assay results but was misdiagnosed by both computed tomography and consistently negative galactomannan assay results. For the galactomannan enzyme-immunoassay the diagnostic odds ratio was 15.33 and for the polymerase chain reaction assay it was 28.67. According to Cohen's kappa our in-house polymerase chain reaction method showed a fair agreement with the galactomannan immunoassay. Combined analysis of the results from the Aspergillus galactomannan enzyme immunoassay together with those generated by our polymerase chain reaction assay led to no misdiagnoses in the control group.
Conclusion:
The data from this pilot-study demonstrate that the consideration of standard clinical methods combined with biomarker testing improves the capacity to make early and more accurate diagnostic decisions.
Background
In this study, we evaluated electrooculography (EOG), an eye tracker and an auditory brain-computer interface (BCI) as access methods to augmentative and alternative communication (AAC). The participant of the study has been in the locked-in state (LIS) for 6 years due to amyotrophic lateral sclerosis. He was able to communicate with slow residual eye movements, but had no means of partner independent communication. We discuss the usability of all tested access methods and the prospects of using BCIs as an assistive technology.
Methods
Within four days, we tested whether EOG, eye tracking and a BCI would allow the participant in LIS to make simple selections. We optimized the parameters in an iterative procedure for all systems.
Results
The participant was able to gain control over all three systems. Nonetheless, due to the level of proficiency previously achieved with his low-tech AAC method, he did not consider using any of the tested systems as an additional communication channel. However, he would consider using the BCI once control over his eye muscles would no longer be possible. He rated the ease of use of the BCI as the highest among the tested systems, because no precise eye movements were required; but also as the most tiring, due to the high level of attention needed to operate the BCI.
Conclusions
In this case study, the partner based communication was possible due to the good care provided and the proficiency achieved by the interlocutors. To ease the transition from a low-tech AAC method to a BCI once control over all muscles is lost, it must be simple to operate. For persons, who rely on AAC and are affected by a progressive neuromuscular disease, we argue that a complementary approach, combining BCIs and standard assistive technology, can prove valuable to achieve partner independent communication and ease the transition to a purely BCI based approach. Finally, we provide further evidence for the importance of a user-centered approach in the design of new assistive devices.
Measurements of the ZZ and WW final states in the mass range above the 2m\(_Z\) and 2m\(_W\) thresholds provide a unique opportunity to measure the off-shell coupling strength of the Higgs boson. This paper presents constraints on the off-shell Higgs boson event yields normalised to the Standard Model prediction (signal strength) in the ZZ→4ℓ, ZZ→2ℓ2ν and WW→eνμν final states. The result is based on pp collision data collected by the ATLAS experiment at the LHC, corresponding to an integrated luminosity of 20.3 fb\(^{-1}\) at a collision energy of \(\sqrt {s}\)=8 TeV. Using the CL\(_S\) method, the observed 95 % confidence level (CL) upper limit on the off-shell signal strength is in the range 5.1–8.6, with an expected range of 6.7–11.0. In each case the range is determined by varying the unknown gg→ZZ and gg→WW background K-factor from higher-order quantum chromodynamics corrections between half and twice the value of the known signal K-factor. Assuming the relevant Higgs boson couplings are independent of the energy scale of the Higgs boson production, a combination with the on-shell measurements yields an observed (expected) 95 % CL upper limit on Γ\(_H\)/Γ\(^{SM}_{H}\) in the range 4.5–7.5 (6.5–11.2) using the same variations of the background K-factor. Assuming that the unknown gg→VV background K-factor is equal to the signal K-factor, this translates into an observed (expected) 95 % CL upper limit on the Higgs boson total width of 22.7 (33.0) MeV.