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Background: Hypothermia has been discussed as playing a role in improving the early phase of systemic inflammation. However, information on the impact of hypothermia on the local inflammatory response is sparse. We therefore investigated the kinetics of local and systemic inflammation in the late posttraumatic phase after induction of hypothermia in an established porcine long-term model of combined trauma.
Materials & Methods: Male pigs (35 ± 5kg) were mechanically ventilated and monitored over the study period of 48 h. Combined trauma included tibia fracture, lung contusion, liver laceration and pressure-controlled hemorrhagic shock (MAP < 30 ± 5 mmHg for 90 min). After resuscitation, hypothermia (33°C) was induced for a period of 12 h (HT-T group) with subsequent re-warming over a period of 10 h. The NT-T group was kept normothermic. Systemic and local (fracture hematoma) cytokine levels (IL-6, -8, -10) and alarmins (HMGB1, HSP70) were measured via ELISA.
Results: Severe signs of shock as well as systemic and local increases of pro-inflammatory mediators were observed in both trauma groups. In general the local increase of pro- and anti-inflammatory mediator levels was significantly higher and prolonged compared to systemic concentrations. Induction of hypothermia resulted in a significantly prolonged elevation of both systemic and local HMGB1 levels at 48 h compared to the NT-T group. Correspondingly, local IL-6 levels demonstrated a significantly prolonged increase in the HT-T group at 48 h.
Conclusion: A prolonged inflammatory response might reduce the well-described protective effects on organ and immune function observed in the early phase after hypothermia induction. Furthermore, local immune response also seems to be affected. Future studies should aim to investigate the use of therapeutic hypothermia at different degrees and duration of application.
Background: Common ECG criteria such as ST-segment changes are of limited value in patients with suspected acute myocardial infarction (AMI) and bundle branch block or wide QRS complex. A large proportion of these patients do not suffer from an AMI, whereas those with ST-elevation myocardial infarction (STEMI) equivalent AMI benefit from an aggressive treatment. Aim of the present study was to evaluate the diagnostic information of cardiac troponin I (cTnI) in hemodynamically stable patients with wide QRS complex and suspected AMI.
Methods: In 417 out of 1818 patients presenting consecutively between 01/2007 and 12/2008 in a prospective multicenter observational study with suspected AMI a prolonged QRS duration was observed. Of these, n = 117 showed significant obstructive coronary artery disease (CAD) used as diagnostic outcome variable. cTnI was determined at admission.
Results: Patients with significant CAD had higher cTnI levels compared to individuals without (median 250ng/L vs. 11ng/L; p<0.01). To identify patients needing a coronary intervention, cTnI yielded an area under the receiver operator characteristics curve of 0.849. Optimized cut-offs with respect to a sensitivity driven rule-out and specificity driven rule-in strategy were established (40ng/L/96ng/L). Application of the specificity optimized cut-off value led to a positive predictive value of 71% compared to 59% if using the 99th percentile cut-off. The sensitivity optimized cut-off value was associated with a negative predictive value of 93% compared to 89% provided by application of the 99th percentile threshold.
Conclusion: cTnI determined in hemodynamically stable patients with suspected AMI and wide QRS complex using optimized diagnostic thresholds improves rule-in and rule-out with respect to presence of a significant obstructive CAD.
Background: Advanced liver diseases are associated with profound alterations of the coagulation system increasing the risk not only of bleeding, but also of thromboembolic complications. A recent milestone study has shown that prophylactic anticoagulation in liver cirrhosis patients results in a reduced frequency of hepatic decompensation. Yet, INR measurement, one of the most widely applied tests to assess liver function, only inaccurately predicts the risk of hepatic decompensation related to alterations of the coagulation system. To assess the relationship between selected coagulation factors / natural anticoagulants with INR, MELD score, and hepatic decompensation, we performed the present pilot study. A total number of 92 patients with various stages of liver cirrhosis were included and prospectively followed for at least 6 months. We found that important natural anticoagulants, namely antithrombin and protein C, as well as factor XI (which may also serve as an anticoagulant) decreased earlier and by a larger magnitude than one would expect from classical coagulation test results. The correlation between these factors and INR was only moderate. Importantly, reduced plasma activities of natural anticoagulants but not INR or MELD score were independent predictors of hepatic encephalopathy (P = 0.013 and 0.003 for antithrombin and protein C, respectively).
Conclusion: In patients with liver cirrhosis plasma activities of several natural anticoagulants are earlier and stronger affected than routine coagulation tests. Reduced activities of natural anticoagulants may be predictive for the development of hepatic encephalopathy.
Thromboembolic events are one of the world’s leading causes of death among patients. Embolus or clot formations have several etiologies including paraneoplastic, post-surgery, cauterization, transplantation, or extracorporeal circuits. Despite its medical significance, little progress has been made in early embolus detection, screening and control. The aim of our study is to test the utility of the in vivo photoacoustic (PA) flow cytometry (PAFC) technique for non-invasive embolus detection in real-time. Using in vivo PAFC, emboli were non-invasively monitored in the bloodstream of two different mouse models. The tumor-free mouse model consisted of two groups, one in which the limbs were clamped to produce vessel stasis (7 procedures), and one where the mice underwent surgery (7 procedures). The melanoma-bearing mouse model also consisted of two groups, one in which the implanted tumor underwent compression (8 procedures), and one where a surgical excision of the implanted tumor was performed (8 procedures). We demonstrated that the PAFC can detect a single embolus, and has the ability to distinguish between erythrocyte–rich (red) and leukocyte/platelet-rich (white) emboli in small vessels. We show that, in tumor-bearing mice, the level of circulating emboli was increased compared to tumor-free mice (p = 0.0013). The number of circulating emboli temporarily increased in the tumor-free control mice during vessel stasis (p = 0.033) and after surgical excisions (signed-rank p = 0.031). Similar observations were noted during tumor compression (p = 0.013) and after tumor excisions (p = 0.012). For the first time, it was possible to detect unlabeled emboli in vivo non-invasively, and to confirm the presence of pigmented tumor cells within circulating emboli. The insight on embolus dynamics during cancer progression and medical procedures highlight the clinical potential of PAFC for early detection of cancer and surgery-induced emboli to prevent the fatal thromboembolic complications by well-timed therapy.
Leptomeningeal dissemination of a primary brain tumor is a condition which is challenging to treat, as it often occurs in rather late disease stages in highly pretreated patients. Its prognosis is dismal and there is still no accepted standard of care. We report here a good clinical effect with a partial response in three out of nine patients and a stable disease with improvement on symptoms in two more patients following systemic anti-angiogenic treatment with bevacizumab (BEV) alone or in combination with chemo- and/or radiotherapy in a series of patients with leptomeningeal dissemination from primary brain tumors (diffuse astrocytoma WHO°II, anaplastic astrocytoma WHO°III, anaplastic oligodendroglioma WHO°III, primitive neuroectodermal tumor and glioblastoma, both WHO°IV). This translated into effective symptom control in five out of nine patients, but only moderate progression-free and overall survival times were reached. Partial responses as assessed by RANO criteria were observed in three patients (each one with anaplastic oligodendroglioma, primitive neuroectodermal tumor and glioblastoma). In these patients progression-free survival (PFS) intervals of 17, 10 and 20 weeks were achieved. In three patients (each one with diffuse astrocytoma, anaplastic astrocytoma and primitive neuroectodermal tumor) stable disease was observed with PFS of 13, 30 and 8 weeks. Another three patients (all with glioblastoma) were primary non-responders and deteriorated rapidly with PFS of 3 to 4 weeks. No severe adverse events were seen. These experiences suggest that the combination of BEV with more conventional therapy schemes with chemo- and/or radiotherapy may be a palliative treatment option for patients with leptomeningeal dissemination of brain tumors.
Triple therapy of chronic hepatitis C virus (HCV) infection with boceprevir (BOC) or telaprevir (TVR) leads to virologic failure in many patients which is often associated with the selection of resistance-associated variants (RAVs). These resistance profiles are of importance for the selection of potential rescue treatment options. In this study, we sequenced baseline NS3 RAVs population-based and investigated the sensitivity of NS3 phenotypes in an HCV replicon assay together with clinical factors for a prediction of treatment response in a cohort of 165 German and Swiss patients treated with a BOC or TVR-based triple therapy. Overall, the prevalence of baseline RAVs was low, although the frequency of RAVs was higher in patients with virologic failure compared to those who achieved a sustained virologic response (SVR) (7% versus 1%, P = 0.06). The occurrence of RAVs was associated with a resistant NS3 quasispecies phenotype (P<0.001), but the sensitivity of phenotypes was not associated with treatment outcome (P = 0.2). The majority of single viral and host predictors of SVR was only weakly associated with treatment response. In multivariate analyses, low AST levels, female sex and an IFNL4 CC genotype were independently associated with SVR. However, a combined analysis of negative predictors revealed a significantly lower overall number of negative predictors in patients with SVR in comparison to individuals with virologic failure (P<0.0001) and the presence of 2 or less negative predictors was indicative for SVR. These results demonstrate that most single baseline viral and host parameters have a weak influence on the response to triple therapy, whereas the overall number of negative predictors has a high predictive value for SVR.
Background: Expected growth in the demand for health services has generated interest in the more effective deployment of health care assistants. Programs encouraging German general practitioners (GPs) to share responsibility for care with specially qualified health care assistants in the family practice (VERAHs) have existed for several years. But no studies have been conducted on the tasks German GPs are willing to rely on specially qualified personnel to perform, what they are prepared to delegate to all non-physician practice staff and what they prefer to do themselves.
Methods: As part of an evaluation study on the deployment of VERAHs in GP-centered health care, we used a questionnaire to ask about task delegation within the practice team. From a list of tasks that VERAHs are specifically trained to carry out, GPs were asked to indicate which they actually delegate. We also asked GPs why they had employed a VERAH in their practice and for their opinions on the benefits and limitations of assigning tasks to VERAHs. The aim of the study was to find out which tasks GPs delegate to their specially qualified personnel, which they permit all HCAs to carry out, and which tasks they do not delegate at all.
Results: The survey was filled in and returned by 245 GPs (83%). Some tasks were exclusively delegated to VERAHs (e.g. home visits), while others were delegated to all HCAs (e.g. vaccinations). About half the GPs rated the assessment of mental health, as part of the comprehensive assessment of a patient’s condition, as the sole responsibility of a GP.
The possibility to delegate more complex tasks was the main reason given for employing a VERAH. Doctors said the delegation of home visits provided them with the greatest relief.
Conclusions: In Germany, where GPs are solely accountable for the health care provided in their practices, experience with the transfer of responsibility to other non-physician health care personnel is still very limited. When HCAs have undergone special training, GPs seem to be prepared to delegate tasks that demand a substantial degree of know-how, such as home visits and case management. This “new” role allocation within the practice may signal a shift in the provision of health care by family practice teams in Germany.
Objective: Mucoactive drugs should increase the ability to expectorate sputum and, ideally, have anti-inflammatory properties. The aim of the study was to evaluate the mucolytic activity of Tyloxapol compared to saline (0.9%) in COPD.
Design: A randomized, placebo-controlled, double-blinded crossover, clinical trial was carried out. Patients were randomly assigned to either inhale 5 ml Tyloxapol 1% or saline 0.9% solution three times daily for 3 weeks and vice versa for another 3 weeks. 28 patients (18 male, 10 female, 47 to 73 years old, median age 63.50) were screened, 21 were treated and 19 patients completed the study per protocol.
Results: A comparison of the two treatment phases showed that the primary endpoint sputum weight was statistically significant higher when patients inhaled Tyloxapol (mean 4.03 g, 95% CI: 2.34–5.73 g at week 3) compared to saline (mean 2.63 g, 95% CI: 1.73–3.53 g at week 3). The p-value at three weeks of treatment was 0.041 between treatment arms. Sputum cells decreased during the Tyloxapol treatment after 3 weeks, indicating that Tyloxapol might have some anti-neutrophilic properties. Lung function parameters (FVC, FEV1, RV, and RV/TLC) remained stable during the study, and no treatment effect was shown. Interestingly, there was a mean increase in all inflammatory cytokines (IL-1β, IL-6, and IL-8) during the saline treatment from day 1 to week 3, whereas during the Tyloxapol treatment, all cytokines decreased. Due to the small sample size and the large individual variation in sputum cytokines, these differences were not significant. However, analyses confirmed that Tyloxapol has significant anti-inflammatory properties in vitro. Despite the high number of inhalations (more than 1000), only 27 adverse events (20 during the Tyloxapol and seven during saline) were recorded. Eleven patients experienced AEs under Tyloxapol and six under saline treatment, which indicates that inhalation of saline or Tyloxapol is a very safe procedure.
Conclusion: Our study demonstrated that inhalation of Tyloxapol by patients with COPD is safe and superior to saline and has some anti-inflammatory effects.
We integrated recent improvements within the floating catchment area (FCA) method family into an integrated ‘iFCA`method. Within this method we focused on the distance decay function and its parameter. So far only distance decay functions with constant parameters have been applied. Therefore, we developed a variable distance decay function to be used within the FCA method. We were able to replace the impedance coefficient β by readily available distribution parameter (i.e. median and standard deviation (SD)) within a logistic based distance decay function. Hence, the function is shaped individually for every single population location by the median and SD of all population-to-provider distances within a global catchment size. Theoretical application of the variable distance decay function showed conceptually sound results. Furthermore, the existence of effective variable catchment sizes defined by the asymptotic approach to zero of the distance decay function was revealed, satisfying the need for variable catchment sizes. The application of the iFCA method within an urban case study in Berlin (Germany) confirmed the theoretical fit of the suggested method. In summary, we introduced for the first time, a variable distance decay function within an integrated FCA method. This function accounts for individual travel behaviors determined by the distribution of providers. Additionally, the function inherits effective variable catchment sizes and therefore obviates the need for determining variable catchment sizes separately.
Background: Dengue virus infection is the most rapidly spreading vector-borne disease in the world. Essential research on dengue virus transmission and its prevention requires community participation. Therefore, it is crucial to understand the factors that are associated with the willingness of communities in high prevalence areas to participate in dengue research. The aim of this study was to explore factors associated with the willingness of healthy community members in Aceh province, Indonesia, to participate in dengue research that would require phlebotomy.
Methodology/Principal Findings: A community-based cross-sectional study was carried out in nine regencies and municipalities of Aceh from November 2014 to March 2015. Interviews using a set of validated questionnaires were conducted to collect data on demography, history of dengue infection, socioeconomic status, and knowledge, attitude and practice regarding dengue fever. Two-step logistic regression and Spearman’s rank correlation (rs) analysis were used to assess the influence of independent variables on dependent variables. Among 535 participants, less than 20% had a good willingness to participate in the dengue study. The factors associated with good willingness to participate were being female, working as a civil servant, private employee or entrepreneur, having a high socioeconomic status and good knowledge, attitude and practice regarding dengue. Good knowledge and attitude regarding dengue were positive independent predictors of willingness to participate (OR: 2.30 [95% CI: 1.36–3.90] and 3.73 [95% CI: 2.24–6.21], respectively).
Conclusion/Significance: The willingness to participate in dengue research is very low among community members in Aceh, and the two most important associated factors are knowledge and attitude regarding dengue. To increase participation rate, efforts to improve the knowledge and attitude of community members regarding dengue fever and dengue-related research is required before such studies are launched.
1. Objective: Chronic hepatitis C virus infections (HCV) cause a significant public health burden. Introduction of telaprevir (TVR) and boceprevir (BOC) has increased sustained virologic response rates (SVR) in genotype 1 patients but were accompanied by higher treatment costs and more side effects. Aim of the study was to assess outcomes and costs of treating HCV with TVR or BOC in routine care.
2. Material and Methods: Data was obtained from a non-interventional study. This analysis relates on a subset of 1,786 patients for whom resource utilisation was documented. Sociodemografic and clinical parameters as well as resource utilisation were collected using a web-based data recording system. Costs were calculated using official remuneration schemes.
3. Results: Mean age of patients was 49.2 years, 58.6% were male. In treatment-naive patients SVR-rates of 62.2% and 55.7% for TVR and BOC were observed (prior relapser: 68.5% for TVR and 63.5% for BOC; prior nonresponder: 45.6% for TVR and 39.1% for BOC). Treatment costs are dominated by costs for pharmaceuticals and range between €39,081 and €53,491. We calculated average costs per SVR of €81,347 (TVR) and €70,163 (BOC) in treatment-naive patients (prior relapser: 78,089 €/SVR for TVR and 82,077 €/SVR for BOC; prior non-responder: 116,509 €/SVR for TVR and 110,156 €/SVR for BOC). Quality of life data showed a considerable decrease during treatment.
4. Conclusion: Our study is one of few investigating both, outcomes and costs, of treating HCV in a real-life setting. Data can serve as a reference in the discussion of increasing costs in recently introduced agents
Mitochondrial cristae are connected to the inner boundary membrane via crista junctions which are implicated in the regulation of oxidative phosphorylation, apoptosis, and import of lipids and proteins. The MICOS complex determines formation of crista junctions. We performed complexome profiling and identified Mic13, also termed Qil1, as a subunit of the MICOS complex. We show that MIC13 is an inner membrane protein physically interacting with MIC60, a central subunit of the MICOS complex. Using the CRISPR/Cas method we generated the first cell line deleted for MIC13. These knockout cells show a complete loss of crista junctions demonstrating that MIC13 is strictly required for the formation of crista junctions. MIC13 is required for the assembly of MIC10, MIC26, and MIC27 into the MICOS complex. However, it is not needed for the formation of the MIC60/MIC19/MIC25 subcomplex suggesting that the latter is not sufficient for crista junction formation. MIC13 is also dispensable for assembly of respiratory chain complexes and for maintaining mitochondrial network morphology. Still, lack of MIC13 resulted in a moderate reduction of mitochondrial respiration. In summary, we show that MIC13 has a fundamental role in crista junction formation and that assembly of respiratory chain supercomplexes is independent of mitochondrial cristae shape.
Background & Aim: The resistance profile of anti-hepatitis C virus (HCV) agents used in combination is important to guide optimal treatment regimens. We evaluated baseline and treatment-emergent NS3/4A and NS5B amino-acid variants among HCV genotype (GT)-1a and -1b-infected patients treated with faldaprevir (HCV protease inhibitor), deleobuvir (HCV polymerase non-nucleoside inhibitor), and ribavirin in multiple clinical studies.
Methods: HCV NS3/4A and NS5B population sequencing (Sanger method) was performed on all baseline plasma samples (n = 1425 NS3; n = 1556 NS5B) and on post-baseline plasma samples from patients with virologic failure (n = 113 GT-1a; n = 221 GT-1b). Persistence and time to loss of resistance-associated variants (RAVs) was estimated using Kaplan–Meier analysis.
Results: Faldaprevir RAVs (NS3 R155 and D168) and deleobuvir RAVs (NS5B 495 and 496) were rare (<1%) at baseline. Virologic response to faldaprevir/deleobuvir/ribavirin was not compromised by common baseline NS3 polymorphisms (e.g. Q80K in 17.5% of GT-1a) or by NS5B A421V, present in 20% of GT-1a. In GT-1b, alanine at NS5B codon 499 (present in 15% of baseline sequences) was associated with reduced response. Treatment-emergent RAVs consolidated previous findings: NS3 R155 and D168 were key faldaprevir RAVs; NS5B A421 and P495 were key deleobuvir RAVs. Among on-treatment virologic breakthroughs, RAVs emerged in both NS3 and NS5B (>90%). Virologic relapse was associated with RAVs in both NS3 and NS5B (53% GT-1b; 52% GT-1b); some virologic relapses had NS3 RAVs only (47% GT-1a; 17% GT-1b). Median time to loss of GT-1b NS5B P495 RAVs post-treatment (5 months) was less than that of GT-1b NS3 D168 (8.5 months) and GT-1a R155 RAVs (11.5 months).
Conclusion: Faldaprevir and deleobuvir RAVs are more prevalent among virologic failures than at baseline. Treatment response was not compromised by common NS3 polymorphisms; however, alanine at NS5B amino acid 499 at baseline (wild-type in GT-1a, polymorphism in GT-1b) may reduce response to this deleobuvir-based regimen.
Introduction: Sepsis remains associated with a high mortality rate. Endotoxin has been shown to influence viscoelastic coagulation parameters, thus suggesting a link between endotoxin levels and the altered coagulation phenotype in septic patients. This study evaluated the effects of systemic polyspecific IgM-enriched immunoglobulin (IgM-IVIg) (Pentaglobin® [Biotest, Dreieich, Germany]) on endotoxin activity (EA), inflammatory markers, viscoelastic and conventional coagulation parameters.
Methods: Patients with severe sepsis were identified by daily screening in a tertiary, academic, surgical ICU. After the inclusion of 15 patients, the application of IgM-IVIg (5 mg/kg/d over three days) was integrated into the unit’s standard operation procedure (SOP) to treat patients with severe sepsis, thereby generating “control” and “IgM-IVIg” groups. EA assays, thrombelastometry (ROTEM®) and impedance aggregometry (Multiplate®) were performed on whole blood. Furthermore, routine laboratory parameters were determined according to unit’s standards.
Results: Data from 26 patients were included. On day 1, EA was significantly decreased in the IgM-IVIg group following 6 and 12 hours of treatment (0.51 ±0.06 vs. 0.26 ±0.07, p<0.05 and 0.51 ±0.06 vs. 0.25 ±0.04, p<0.05) and differed significantly compared with the control group following 6 hours of treatment (0.26 ±0.07 vs. 0.43 ±0.07, p<0.05). The platelet count was significantly higher in the IgM-IVIg group following four days of IgM-IVIg treatment (200/nl ±43 vs. 87/nl ±20, p<0.05). The fibrinogen concentration was significantly lower in the control group on day 2 (311 mg/dl ±37 vs. 475 mg/dl ±47 (p = 0.015)) and day 4 (307 mg/dl ±35 vs. 420 mg/dl ±16 (p = 0.017)). No differences in thrombelastometric or aggregometric measurements, or inflammatory markers (interleukin-6 (IL-6), leukocyte, lipopolysaccharide binding protein (LBP)) were observed.
Conclusion: Treatment with IgM-enriched immunoglobulin attenuates the EA levels in patients with severe sepsis and might have an effect on septic thrombocytopenia and fibrinogen depletion. Viscoelastic, aggregometric or inflammatory parameters were not influenced.
Purpose: Quantitative T2'-mapping detects regional changes of the relation of oxygenated and deoxygenated hemoglobin (Hb) by using their different magnetic properties in gradient echo imaging and might therefore be a surrogate marker of increased oxygen extraction fraction (OEF) in cerebral hypoperfusion. Since elevations of cerebral blood volume (CBV) with consecutive accumulation of Hb might also increase the fraction of deoxygenated Hb and, through this, decrease the T2’-values in these patients we evaluated the relationship between T2’-values and CBV in patients with unilateral high-grade large-artery stenosis.
Materials and Methods Data from 16 patients (13 male, 3 female; mean age 53 years) with unilateral symptomatic or asymptomatic high-grade internal carotid artery (ICA) or middle cerebral artery (MCA) stenosis/occlusion were analyzed. MRI included perfusion-weighted imaging and high-resolution T2’-mapping. Representative relative (r)CBV-values were analyzed in areas of decreased T2’ with different degrees of perfusion delay and compared to corresponding contralateral areas.
Results: No significant elevations in cerebral rCBV were detected within areas with significantly decreased T2’-values. In contrast, rCBV was significantly decreased (p<0.05) in regions with severe perfusion delay and decreased T2’. Furthermore, no significant correlation between T2’- and rCBV-values was found. Conclusions rCBV is not significantly increased in areas of decreased T2’ and in areas of restricted perfusion in patients with unilateral high-grade stenosis. Therefore, T2’ should only be influenced by changes of oxygen metabolism, regarding our patient collective especially by an increase of the OEF. T2’-mapping is suitable to detect altered oxygen consumption in chronic cerebrovascular disease.
Background: Astrocytomas are the most common primary brain tumors distinguished into four histological grades. Molecular analyses of individual astrocytoma grades have revealed detailed insights into genetic, transcriptomic andepigenetic alterations. This provides an excellent basis to identify similarities and differences between astrocytoma grades. Methods: We utilized public omics data of all four astrocytoma grades focusing on pilocytic astrocytomas (PA I), diffuse astrocytomas (AS II), anaplastic astrocytomas (AS III) and glioblastomas (GBM IV) to identify similarities and differences using well-established bioinformatics and systems biology approaches. We further validated the expression and localization of Ang2 involved in angiogenesis using immunohistochemistry. Results: Our analyses show similarities and differences between astrocytoma grades at the level of individual genes, signaling pathways and regulatory networks. We identified many differentially expressed genes that were either exclusively observed in a specific astrocytoma grade or commonly affected in specific subsets of astrocytoma grades in comparison to normal brain. Further, the number of differentially expressed genes generally increased with the astrocytoma grade with one major exception. The cytokine receptor pathway showed nearly the same number of differentially expressed genes in PA I and GBM IV and was further characterized by a significant overlap of commonly altered genes and an exclusive enrichment of overexpressed cancer genes in GBM IV. Additional analyses revealed a strong exclusive overexpression of CX3CL1 (fractalkine) and its receptor CX3CR1 in PA I possibly contributing to the absence of invasive growth. We further found that PA I was significantly associated with the mesenchymal subtype typically observed for very aggressive GBM IV. Expression of endothelial and mesenchymal markers (ANGPT2, CHI3L1) indicated a stronger contribution of the micro-environment to the manifestation of the mesenchymal subtype than the tumor biology itself. We further inferred a transcriptional regulatory network associated with specific expression differences distinguishing PA I from AS II, AS III and GBM IV. Major central transcriptional regulators were involved in brain development, cell cycle control, proliferation, apoptosis, chromatin remodeling or DNA methylation. Many of these regulators showed directly underlying DNA methylation changes in PA I or gene copy number mutations in AS II, AS III and GBM IV. Conclusions: This computational study characterizes similarities and differences between all four astrocytoma grades confirming known and revealing novel insights into astrocytoma biology. Our findings represent a valuable resource for future computational and experimental studies.
Background & Aims: Simeprevir is an oral, once-daily inhibitor of hepatitis c virus (HCV) protease NS3/4A. We investigated the safety and efficacy of simeprevir with peg-interferon α-2a and ribavirin (PR) in a randomized, double-blind, placebo-controlled, phase 3 trial of patients with HCV genotype 1 infection who relapsed after previous interferon-based therapy.
Methods: Patients were assigned randomly (2:1) to groups given simeprevir (150 mg, once daily) and PR (n = 260) or placebo and PR (n = 133) for 12 weeks. Patients then were given PR alone for 12 or 36 weeks (simeprevir group, based on response-guided therapy criteria) or 36 weeks (placebo group).
Results: Simeprevir and PR was significantly superior to placebo and PR; rates of sustained virologic response 12 weeks after planned end of treatment (SVR12) were 79.2% vs 36.1%, respectively (43.8% difference; 95% confidence interval, 34.6–53.0; P < .001). Among patients given simeprevir, 92.7% met the response-guided therapy criteria and were eligible to complete PR at week 24; of these, 83.0% achieved SVR12. HCV RNA was undetectable at week 4 in 77.2% of patients given simeprevir and 3.1% given placebo. On-treatment failure and relapse rates were lower among patients given simeprevir and PR than those given placebo and PR (3.1% vs 27.1%, and 18.5% vs 48.4%, respectively). Patients given simeprevir did not have adverse events beyond those that occurred in patients given PR alone. Most adverse events were grades 1/2; the prevalence of anemia and rash was similar in both groups. Patients in both groups reported similar severity of fatigue and functional impairments during the study, but duration was reduced among patients given simeprevir.
Conclusions: In a phase 3 trial of patients who had relapsed after interferon-based therapy, the addition of simeprevir to PR was generally well tolerated, with an SVR12 rate of 79.2%. Most patients (92.7%) receiving simeprevir were able to shorten therapy to 24 weeks. ClinicalTrials.gov number: NCT01281839.
Cardiac arrhythmias are often associated with mutations in ion channels or other proteins. To enable drug development for distinct arrhythmias, model systems are required that allow implementing patient-specific mutations. We assessed a muscular pump in Caenorhabditis elegans. The pharynx utilizes homologues of most of the ion channels, pumps and transporters defining human cardiac physiology. To yield precise rhythmicity, we optically paced the pharynx using channelrhodopsin-2. We assessed pharynx pumping by extracellular recordings (electropharyngeograms--EPGs), and by a novel video-microscopy based method we developed, which allows analyzing multiple animals simultaneously. Mutations in the L-type VGCC (voltage-gated Ca(2+)-channel) EGL-19 caused prolonged pump duration, as found for analogous mutations in the Cav1.2 channel, associated with long QT syndrome. egl-19 mutations affected ability to pump at high frequency and induced arrhythmicity. The pharyngeal neurons did not influence these effects. We tested whether drugs could ameliorate arrhythmia in the optogenetically paced pharynx. The dihydropyridine analog Nemadipine A prolonged pump duration in wild type, and reduced or prolonged pump duration of distinct egl-19 alleles, thus indicating allele-specific effects. In sum, our model may allow screening of drug candidates affecting specific VGCCs mutations, and permit to better understand the effects of distinct mutations on a macroscopic level.
Responses to recent infectious disease outbreaks, such as to Influenza Pandemic 2009 and the on-going Ebola outbreak in West Africa, reveal the need for new and strengthened approaches to risk communication and governance. The article argues for a fundamental re-conceptualisation of current approaches to risk communication, preparedness planning and response. It calls for a reframing of the way we currently identify and respond to outbreaks around a set of core behaviour-based response patterns. This new model moves away from the current risk communication focus on a plethora of agent-specific threats to five generic response patterns that are based on socially relevant response activities such as 1) controlling vectors, 2) enhancing hygiene, 3) isolation of the sick, 4) protection of the well, and 5) systemic protection of people and their environments. Emphasis is placed on gaining relevant insights into the context specific needs of different communities related to these five patterns. Governance structures are then built and evaluated based on their capacity to collect, communicate, share and prepare the public to take appropriate action related to the five different patterns before, during and after an event. Reframing risk communication and preparedness approaches around a better understanding of the determinants of these general behavioural patterns in infectious control could strengthen infection control literacy, response competence and build resilience of both individuals and health systems to address future epidemics, pandemics and other public health threats.
BACKGROUND: Human SAMHD1 is a triphosphohydrolase that restricts the replication of retroviruses, retroelements and DNA viruses in noncycling cells. While modes of action have been extensively described for human SAMHD1, only little is known about the regulation of SAMHD1 in the mouse. Here, we characterize the antiviral activity of murine SAMHD1 with the help of knockout mice to shed light on the regulation and the mechanism of the SAMHD1 restriction and to validate the SAMHD1 knockout mouse model for the use in future infectivity studies.
RESULTS: We found that endogenous mouse SAMHD1 restricts not only HIV-1 but also MLV reporter virus infection at the level of reverse transcription in primary myeloid cells. Similar to the human protein, the antiviral activity of murine SAMHD1 is regulated through phosphorylation at threonine 603 and is limited to nondividing cells. Comparing the susceptibility to infection with intracellular dNTP levels and SAMHD1 phosphorylation in different cell types shows that both functions are important determinants of the antiviral activity of murine SAMHD1. In contrast, we found the proposed RNase activity of SAMHD1 to be less important and could not detect any effect of mouse or human SAMHD1 on the level of incoming viral RNA.
CONCLUSION: Our findings show that SAMHD1 in the mouse blocks retroviral infection at the level of reverse transcription and is regulated through cell cycle-dependent phosphorylation. We show that the antiviral restriction mediated by murine SAMHD1 is mechanistically similar to what is known for the human protein, making the SAMHD1 knockout mouse model a valuable tool to characterize the influence of SAMHD1 on the replication of different viruses in vivo.
Gallbladder cancer (GBC) is a highly malignant tumor characterized by a poor response to chemotherapy and radiotherapy. We evaluated the in vitro and in vivo antitumor efficacy of mTOR inhibitors, rapamycin and WYE-354. In vitro assays showed WYE-354 significantly reduced cell viability, migration and invasion and phospho-P70S6K expression in GBC cells. Mice harboring subcutaneous gallbladder tumors, treated with WYE-354 or rapamycin, exhibited a significant reduction in tumor mass. A short-term treatment with a higher dose of WYE-354 decreased the tumor size by 68.6% and 52.4%, in mice harboring G-415 or TGBC-2TKB tumors, respectively, compared to the control group. By contrast, treatment with a prolonged-low-dose regime of rapamycin almost abrogated tumor growth, exhibiting 92.7% and 97.1% reduction in tumor size, respectively, compared to control mice. These results were accompanied by a greater decrease in the phosphorylation status of P70S6K and a lower cell proliferation Ki67 index, compared to WYE-354 treated mice, suggesting a more effective mTOR pathway inhibition. These findings provide a proof of concept for the use of rapamycin or WYE-354 as potentially good candidates to be studied in clinical trials in GBC patients.
BACKGROUND: Toxoplasmosis endangers the unborn child if its infectious agent--toxoplasma gondii--is transmitted transplacentally during pregnancy. Although this condition occurs in all parts of the world and represents a major public health burden, no detailed knowledge on the global research architecture of congenital toxoplasmosis is available thus far. Hence, it was the aim of this study to assess the related global research activity over the past 110 years.
METHODS: We employed the NewQIS platform, which combines established scientometric and socioeconomic analysis tools with novel visualizing techniques such as density equalizing mapping projections.
RESULTS: In the Web of Science, 13,044 congenital toxoplasmosis-related items published between 1900 and 2012 were identified. These were issued by 26,483 authors originating from 125 countries. The US was the dominating nation (38.7 % of total scientific output), followed by France (10.9 %) and Great Britain (9.2 %). The US also led the ranking in regards to semi-qualitative parameters (total citations, country-specific h-indices and citation rates). When research activity was related to economic figures, the ratio of total toxoplasmosis publications to the total GDP listed Switzerland first with an average of 589.35 toxoplasmosis-related publications per GDP in 1000 Bio US-$, followed by France (545.16), the UK (486.13) and Brazil (431.84) and the US (311.11). The relation of toxoplasmosis-specific publications to the economic power indicator GDP per capita in 1000 US-$ revealed that the US was ranked first with 97.65 toxoplasmosis-related publications/GDP per capita in 1000 US-$, followed by Brazil (85.95). Subject area analysis indicated a relative shortage of studies that addressed pharmacological or public health aspects of congenital toxoplasmosis.
CONCLUSIONS: This study is the first in-depth approach to sketch a global picture of the congenital toxoplasmosis research architecture. In contrast to other fields of biomedical research, not only high-income countries play a major role regarding congenital toxoplasmosis research but also countries such as Brazil that have a high incidence of congenital toxoplasmosis.
Breast cancer is fast becoming the leading cause of oncologic morbidity and mortality among women worldwide. Demographic changes in Asia, Southeast Asia, and South America will further accelerate this trend. Different specialties are involved in the treatment of breast cancer patients: gynecology, surgery, pathology, hematology/oncology, radiology, radiation oncology, and nuclear medicine. Optimal results are seen in countries providing standardized breast cancer care in certified breast centers. The present article provides an overview of current state-of-the-art treatment strategies and explains the contributions of different specialties to optimal and individualized care for breast cancer patients. Breast cancer will be one of the most important health issues facing physicians involved with women’s health and a basic understanding of current treatment objectives will be essential medical knowledge for everyone taking care of female patients.
Nuclear export factor 1 (NXF1) exports mRNA to the cytoplasm after recruitment to mRNA by specific adaptor proteins. How and why cells use numerous different export adaptors is poorly understood. Here we critically evaluate members of the SR protein family (SRSF1-7) for their potential to act as NXF1 adaptors that couple pre-mRNA processing to mRNA export. Consistent with this proposal, >1000 endogenous mRNAs required individual SR proteins for nuclear export in vivo. To address the mechanism, transcriptome-wide RNA-binding profiles of NXF1 and SRSF1-7 were determined in parallel by individual-nucleotide-resolution UV cross-linking and immunoprecipitation (iCLIP). Quantitative comparisons of RNA-binding sites showed that NXF1 and SR proteins bind mRNA targets at adjacent sites, indicative of cobinding. SRSF3 emerged as the most potent NXF1 adaptor, conferring sequence specificity to RNA binding by NXF1 in last exons. Interestingly, SRSF3 and SRSF7 were shown to bind different sites in last exons and regulate 3' untranslated region length in an opposing manner. Both SRSF3 and SRSF7 promoted NXF1 recruitment to mRNA. Thus, SRSF3 and SRSF7 couple alternative splicing and polyadenylation to NXF1-mediated mRNA export, thereby controlling the cytoplasmic abundance of transcripts with alternative 3' ends.
An increasing body of evidences from preclinical as well as epidemiological and clinical studies suggest a potential beneficial role of dietary intake of omega-3 fatty acids for cognitive functioning. In this narrative review, we will summarize and discuss recent findings from epidemiological, interventional and experimental studies linking dietary consumption of omega-3 fatty acids to cognitive function in healthy adults. Furthermore, affective disorders and schizophrenia (SZ) are characterized by cognitive dysfunction encompassing several domains. Cognitive dysfunction is closely related to impaired functioning and quality of life across these conditions. Therefore, the current review focues on the potential influence of omega-3 fatty acids on cognition in SZ and affective disorders. In sum, current data predominantly from mechanistic models and animal studies suggest that adjunctive omega-3 fatty acid supplementation could lead to improved cognitive functioning in SZ and affective disorders. However, besides its translational promise, evidence for clinical benefits in humans has been mixed. Notwithstanding evidences indicate that adjunctive omega-3 fatty acids may have benefit for affective symptoms in both unipolar and bipolar depression, to date no randomized controlled trial had evaluated omega-3 as cognitive enhancer for mood disorders, while a single published controlled trial suggested no therapeutic benefit for cognitive improvement in SZ. Considering the pleiotropic mechanisms of action of omega-3 fatty acids, the design of well-designed controlled trials of omega-3 supplementation as a novel, domain-specific, target for cognitive impairment in SZ and affective disorders is warranted.
The current Review article provides a narrative review about the neurobiological underpinnings and treatment of treatment resistant late-life depression (TRLLD). The manuscript focuses on therapeutic targets of late-life depression, which include pharmacological, psychological, biophysical and exercise treatment approaches. Therefore, we summarize available evidences on that kind of therapies for patients suffering from late-life depression. The search for evidences of therapeutic options of late-life depression were done using searching websites as “pubmed”, and using the searching terms “depression”, “late-life depression”, “treatment”, “biophysical therapy”, “exercise therapy”, “pharmacological therapy” and “psychological therapy”. To the end, we summarize and discuss current data, providing some directions for further research.
Treatment recommendations for elderly depressive patients favour a multimodal approach, containing psychological, pharmacological and secondary biophysical therapeutic options. Particularly, a combination of psychotherapy and antidepressant medication reflects the best therapeutic option. However, mostly accepted and used is the pharmacological treatment although evidence suggests that the drug therapy is not as effective as it is in younger depressive patients. Further studies employing larger samples and longer follow-up periods are necessary and may focus on comparability of study designs and involve novel approaches to establish the validity and reliability of multimodal treatment programs.
Understanding how to achieve efficient transduction of hematopoietic stem and progenitor cells (HSPCs), while preserving their long-term ability to self-reproduce, is key for applying lentiviral-based gene engineering methods. SAMHD1 is an HIV-1 restriction factor in myeloid and resting CD4+ T cells that interferes with reverse transcription by decreasing the nucleotide pools or by its RNase activity. Here we show that SAMHD1 is expressed at high levels in HSPCs cultured in a medium enriched with cytokines. Thus, we hypothesized that degrading SAMHD1 in HSPCs would result in more efficient lentiviral transduction rates. We used viral like particles (VLPs) containing Vpx, shRNA against SAMHD1, or provided an excess of dNTPs or dNs to study this question. Regardless of the method applied, we saw no increase in the lentiviral transduction rate. The result was different when we used viruses (HR-GFP-Vpx+) which carry Vpx and encode GFP. These viruses allow assessment of the effects of Vpx specifically in the transduced cells. Using HR-GFP-Vpx+ viruses, we observed a modest but significant increase in the transduction efficiency. These data suggest that SAMHD1 has some limited efficacy in blocking reverse transcription but the major barrier for efficient lentiviral transduction occurs before reverse transcription.
Despite multimodal regimens and diverse treatment options alleviating disease symptoms, morbidity and mortality associated with advanced ischemic heart failure remain high. Recently, technological innovation has led to the development of regenerative therapeutic interventions aimed at halting or reversing the vicious cycle of heart failure progression. Driven by the unmet patient need and fueled by encouraging experimental studies, stem cell-based clinical trials have been launched over the past decade. Collectively, these trials have enrolled several thousand patients and demonstrated the clinical feasibility and safety of cell-based interventions. However, the totality of evidence supporting their efficacy in ischemic heart failure remains limited. Experience from the early randomized stem cell clinical trials underscores the key points in trial design ranging from adequate hypothesis formulation to selection of the optimal patient population, cell type and delivery route. Importantly, to translate the unprecedented promise of regenerative biotherapies into clinical benefit, it is crucial to ensure the appropriate choice of endpoints along the regulatory path. Accordingly, we here provide considerations relevant to the choice of endpoints for regenerative clinical trials in the ischemic heart failure setting.
As current classical Hodgkin lymphoma (cHL) treatment strategies have pronounced side-effects, specific inhibition of signaling pathways may offer novel strategies in cHL therapy. Basal autophagy, a regulated catabolic pathway to degrade cell's own components, is in cancer linked with both, tumor suppression or promotion. The finding that basal autophagy enhances tumor cell survival would thus lead to immediately testable strategies for novel therapies. Thus, we studied its contribution in cHL.We found constitutive activation of autophagy in cHL cell lines and primary tissue. The expression of key autophagy-relevant proteins (e.g. Beclin-1, ULK1) and LC3 processing was increased in cHL cells, even in lymphoma cases. Consistently, cHL cells exhibited elevated numbers of autophagic vacuoles and intact autophagic flux. Autophagy inhibition with chloroquine or inactivation of ATG5 induced apoptosis and reduced proliferation of cHL cells. Chloroquine-mediated inhibition of basal autophagy significantly impaired HL growth in-vivo in NOD SCID γc-/- (NSG) mice. We found that basal autophagy plays a pivotal role in sustaining mitochondrial function.We conclude that cHL cells require basal autophagy for growth, survival and sustained metabolism making them sensitive to autophagy inhibition. This suggests basal autophagy as useful target for new strategies in cHL treatment.
The aim of this preliminary prospective RCT was to histologically evaluate peri-implant soft tissues around titanium abutments treated using different cleaning methods. Sixteen patients were randomized into three groups: laboratory customized abutments underwent Plasma of Argon treatment (Plasma Group), laboratory customized abutments underwent cleaning by steam (Steam Group), and abutments were used as they came from industry (Control Group). Seven days after the second surgery, soft tissues around abutments were harvested. Samples were histologically analyzed. Soft tissues surrounding Plasma Group abutments predominantly showed diffuse chronic infiltrate, almost no acute infiltrate, with presence of few polymorphonuclear neutrophil granulocytes, and a diffuse presence of collagenization bands. Similarly, in Steam Group, the histological analysis showed a high variability of inflammatory expression factors. Tissues harvested from Control Group showed presence of few neutrophil granulocytes, moderate presence of lymphocytes, and diffuse collagenization bands in some sections, while they showed absence of acute infiltrate in 40% of sections. However, no statistical difference was found among the tested groups for each parameter (p > 0.05). Within the limit of the present study, results showed no statistically significant difference concerning inflammation and healing tendency between test and control groups.
OBJECTIVES: Outcome of aortic valve replacement may be influenced by the choice of bioprosthesis. Pericardial heart valves are described to have a favourable haemodynamic profile compared with porcine valves, although the clinical notability of this finding is still controversially debated. Herein, we compared the long-term results of two commonly implanted bioprosthesis at a single centre.
METHODS: All consecutive patients undergoing isolated aortic valve replacement with either a Carpentier-Edwards Magna pericardial prosthesis or a Medtronic Mosaic porcine prosthesis between 2002 and 2008 were analysed regarding preoperative characteristics, short- and long-term survival, valve-related complications and echocardiographic findings.
RESULTS: The Medtronic Mosaic was implanted in 163 patients and the Carpentier-Edwards Magna in 295 patients. The sizes of implanted valves were 22.4 ± 1.5 mm for the Mosaic and 21.8 ± 1.8 mm for the Magna (P = 0.001). The long-term survival rate was 76 and 56% after 5 and 10 years for the Medtronic Mosaic, which was comparable with the Carpentier-Edwards Magna (77 and 57%; P = 0.92). Overall long-term survival was comparable with an age- and sex-matched Austrian general population for both groups. Valve-related adverse events were similar between groups. The postoperative mean transvalvular gradient was significantly increased in the Mosaic group (24 ± 9 mmHg vs 17 ± 7 mmHg; P < 0.001).
CONCLUSIONS: Both types of aortic bioprostheses offer excellent results after isolated aortic valve replacement. Despite relevant differences in gradients, long-term survival was comparable with the expected normal survival for both bioprostheses. Patients with a porcine heart valve had a higher postoperative transvalvular gradient.
The processing of pain undergoes several changes in aging that affect sensory nociceptive fibers and the endogenous neuronal inhibitory systems. So far, it is not completely clear whether age-induced modifications are associated with an increase or decrease in pain perception. In this study, we assessed the impact of age on inflammatory nociception in mice and the role of the hormonal inhibitory systems in this context. We investigated the nociceptive behavior of 12-month-old versus 6–8-week-old mice in two behavioral models of inflammatory nociception. Levels of TRP channels, and cortisol as well as cortisol targets, were measured by qPCR, ELISA, and Western blot in the differently aged mice. We observed an age-related reduction in nociceptive behavior during inflammation as well as a higher level of cortisol in the spinal cord of aged mice compared to young mice, while TRP channels were not reduced. Among potential cortisol targets, the NF-κB inhibitor protein alpha (IκBα) was increased, which might contribute to inhibition of NF-κB and a decreased expression and activity of the inducible nitric oxide synthase (iNOS). In conclusion, our results reveal a reduced nociceptive response in aged mice, which might be at least partially mediated by an augmented inflammation-induced increase in the hormonal inhibitory system involving cortisol.
Objectives: Patient safety is a crucial issue in medicine. Its main objective is to reduce the number of deaths and health damages that are caused by preventable medical errors. To achieve this, it needs better health systems that make mistakes less likely and their effects less detrimental without blaming health workers for failures. Until now, there is no in-depth scientometric analysis on this issue that encompasses the interval between 1963 and 2014. Therefore, the aim of this study is to sketch a landscape of the past global research output on patient safety including the gender distribution of the medical discipline of patient safety by interpreting scientometric parameters. Additionally, respective future trends are to be outlined.
Setting: The Core Collection of the scientific database Web of Science was searched for publications with the search term ‘Patient Safety’ as title word that was focused on the corresponding medical discipline. The resulting data set was analysed by using the methodology implemented by the platform NewQIS. To visualise the geographical landscape, state-of-the-art techniques including density-equalising map projections were applied.
Results: 4079 articles on patient safety were identified in the period from 1900 to 2014. Most articles were published in North America, the UK and Australia. In regard to the overall number of publications, the USA is the leading country, while the output ratio to the population of Switzerland was found to exhibit the best performance. With regard to the ratio of the number of publications to the Gross Domestic Product (GDP) per Capita, the USA remains the leading nation but countries like India and China with a low GDP and high population numbers are also profiting.
Conclusions: Though the topic is a global matter, the scientific output on patient safety is centred mainly in industrialised countries.
CD4+CD25+ regulatory T cells (Tregs) represent a specialized subpopulation of T cells, which are essential for maintaining peripheral tolerance and preventing autoimmunity. The immunomodulatory effects of Tregs depend on their activation status. Here we show that, in contrast to conventional anti-CD4 monoclonal antibodies (mAbs), the humanized CD4-specific monoclonal antibody tregalizumab (BT-061) is able to selectively activate the suppressive properties of Tregs in vitro. BT-061 activates Tregs by binding to CD4 and activation of signaling downstream pathways. The specific functionality of BT-061 may be explained by the recognition of a unique, conformational epitope on domain 2 of the CD4 molecule that is not recognized by other anti-CD4 mAbs. We found that, due to this special epitope binding, BT-061 induces a unique phosphorylation of T-cell receptor complex-associated signaling molecules. This is sufficient to activate the function of Tregs without activating effector T cells. Furthermore, BT-061 does not induce the release of pro-inflammatory cytokines. These results demonstrate that BT-061 stimulation via the CD4 receptor is able to induce T-cell receptor-independent activation of Tregs. Selective activation of Tregs via CD4 is a promising approach for the treatment of autoimmune diseases where insufficient Treg activity has been described. Clinical investigation of this new approach is currently ongoing.
Background Reaching the therapeutic target of remission or low-disease activity has improved outcomes in patients with rheumatoid arthritis (RA) significantly. The treat-to-target recommendations, formulated in 2010, have provided a basis for implementation of a strategic approach towards this therapeutic goal in routine clinical practice, but these recommendations need to be re-evaluated for appropriateness and practicability in the light of new insights.
Objective To update the 2010 treat-to-target recommendations based on systematic literature reviews (SLR) and expert opinion.
Methods A task force of rheumatologists, patients and a nurse specialist assessed the SLR results and evaluated the individual items of the 2010 recommendations accordingly, reformulating many of the items. These were subsequently discussed, amended and voted upon by >40 experts, including 5 patients, from various regions of the world. Levels of evidence, strengths of recommendations and levels of agreement were derived.
Results The update resulted in 4 overarching principles and 10 recommendations. The previous recommendations were partly adapted and their order changed as deemed appropriate in terms of importance in the view of the experts. The SLR had now provided also data for the effectiveness of targeting low-disease activity or remission in established rather than only early disease. The role of comorbidities, including their potential to preclude treatment intensification, was highlighted more strongly than before. The treatment aim was again defined as remission with low-disease activity being an alternative goal especially in patients with long-standing disease. Regular follow-up (every 1–3 months during active disease) with according therapeutic adaptations to reach the desired state was recommended. Follow-up examinations ought to employ composite measures of disease activity that include joint counts. Additional items provide further details for particular aspects of the disease, especially comorbidity and shared decision-making with the patient. Levels of evidence had increased for many items compared with the 2010 recommendations, and levels of agreement were very high for most of the individual recommendations (≥9/10).
Conclusions The 4 overarching principles and 10 recommendations are based on stronger evidence than before and are supposed to inform patients, rheumatologists and other stakeholders about strategies to reach optimal outcomes of RA.
Spinocerebellar ataxia type 2 (SCA2) and amyotrophic lateral sclerosis (ALS) are neurodegenerative disorders, caused or modified by an unstable CAG-repeat expansion in the SCA2 gene, which encodes a polyglutamine (polyQ) domain expansion in ataxin-2 (ATXN2). ATXN2 is an RNA-binding protein and interacts with the poly(A)-binding protein PABPC1, localizing to ribosomes at the rough endoplasmic reticulum. Under cell stress, ATXN2, PABPC1 and small ribosomal subunits are relocated to stress granules, where mRNAs are protected from translation and from degradation. It is unknown whether ATXN2 associates preferentially with specific mRNAs or how it modulates RNA processing. Here, we investigated the RNA profile of the liver and cerebellum from Atxn2 knockout (Atxn2−/−) mice at two adult ages, employing oligonucleotide microarrays. Prominent increases were observed for Lsm12/Paip1 (>2-fold), translation modulators known as protein interactor/competitor of ATXN2 and for Plin3/Mttp (>1.3-fold), known as apolipoprotein modulators in agreement with the hepatosteatosis phenotype of the Atxn2−/− mice. Consistent modest upregulations were also observed for many factors in the ribosome and the translation/secretion apparatus. Quantitative reverse transcriptase PCR in liver tissue validated >1.2-fold upregulations for the ribosomal biogenesis modulator Nop10, the ribosomal components Rps10, Rps18, Rpl14, Rpl18, Gnb2l1, the translation initiation factors Eif2s2, Eif3s6, Eif4b, Pabpc1 and the rER translocase factors Srp14, Ssr1, Sec61b. Quantitative immunoblots substantiated the increased abundance of NOP10, RPS3, RPS6, RPS10, RPS18, GNB2L1 in SDS protein fractions, and of PABPC1. In mouse embryonal fibroblasts, ATXN2 absence also enhanced phosphorylation of the ribosomal protein S6 during growth stimulation, while impairing the rate of overall protein synthesis rates, suggesting a block between the enhanced translation drive and the impaired execution. Thus, the physiological role of ATXN2 subtly modifies the abundance of cellular translation factors as well as global translation.
Mesenchymal stem/stromal cells (MSCs) feature promising potential for cellular therapies, yet significant progress in development of MSC therapeutics and assays is hampered because of remarkable MSC heterogeneity in vivo and in vitro. This heterogeneity poses challenges for standardization of MSC characterization and potency assays as well as for MSC study comparability and manufacturing. This review discusses promising marker combinations for prospective MSC subpopulation enrichment and expansion, and reflects MSC phenotype changes due to environment and age. In order to address animal modelling in MSC biology, comparison of mouse and human MSC markers highlights current common ground of MSCs between species.
Ionizing radiation generates DNA double-strand breaks (DSB) which, unless faithfully repaired, can generate chromosomal rearrangements in hematopoietic stem and/or progenitor cells (HSPC), potentially priming the cells towards a leukemic phenotype. Using an enhanced green fluorescent protein (EGFP)-based reporter system, we recently identified differences in the removal of enzyme-mediated DSB in human HSPC versus mature peripheral blood lymphocytes (PBL), particularly regarding homologous DSB repair (HR). Assessment of chromosomal breaks via premature chromosome condensation or γH2AX foci indicated similar efficiency and kinetics of radiation-induced DSB formation and rejoining in PBL and HSPC. Prolonged persistence of chromosomal breaks was observed for higher LET charged particles which are known to induce more complex DNA damage compared to X-rays. Consistent with HR deficiency in HSPC observed in our previous study, we noticed here pronounced focal accumulation of 53BP1 after X-ray and carbon ion exposure (intermediate LET) in HSPC versus PBL. For higher LET, 53BP1 foci kinetics was similarly delayed in PBL and HSPC suggesting similar failure to repair complex DNA damage. Data obtained with plasmid reporter systems revealed a dose- and LET-dependent HR increase after X-ray, carbon ion and higher LET exposure, particularly in HR-proficient immortalized and primary lymphocytes, confirming preferential use of conservative HR in PBL for intermediate LET damage repair. HR measured adjacent to the leukemia-associated MLL breakpoint cluster sequence in reporter lines revealed dose dependency of potentially leukemogenic rearrangements underscoring the risk of leukemia-induction by radiation treatment.
The inhibitor of the nuclear factor-κB (IκB) kinase (IKK) complex is a key regulator of the canonical NF-κB signalling cascade and is crucial for fundamental cellular functions, including stress and immune responses. The majority of IKK complex functions are attributed to NF-κB activation; however, there is increasing evidence for NF-κB pathway-independent signalling. Here we combine quantitative mass spectrometry with random forest bioinformatics to dissect the TNF-α-IKKβ-induced phosphoproteome in MCF-7 breast cancer cells. In total, we identify over 20,000 phosphorylation sites, of which ∼1% are regulated up on TNF-α stimulation. We identify various potential novel IKKβ substrates including kinases and regulators of cellular trafficking. Moreover, we show that one of the candidates, AEG-1/MTDH/LYRIC, is directly phosphorylated by IKKβ on serine 298. We provide evidence that IKKβ-mediated AEG-1 phosphorylation is essential for IκBα degradation as well as NF-κB-dependent gene expression and cell proliferation, which correlate with cancer patient survival in vivo.
BACKGROUND: Plasminogen deficiency is a rare autosomal recessive disease, which is associated with aggressive periodontitis and gingival enlargement. Previously described treatments of plasminogen deficiency associated periodontitis have shown limited success. This is the first case report indicating a successful therapy approach consisting of a non-surgical supra- and subgingival debridement in combination with an adjunctive systemic antibiotic therapy and a strict supportive periodontal regimen over an observation period of 4 years.
CASE PRESENTATION: The intraoral examination of a 17-year-old Turkish female with severe plasminogen deficiency revealed generalized increased pocket probing depths ranging from 6 to 9 mm, bleeding on probing over 30%, generalized tooth mobility, and gingival hyperplasia. Alveolar bone loss ranged from 30% to 50%. Clinical attachment loss corresponded to pocket probing depths. Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Treponema denticola, Prevotella intermedia, Prevotella nigrescens and Eikenella corrodens have been detected by realtime polymerase chain reaction. Periodontal treatment consisted of full mouth disinfection and adjunctive systemic administration of amoxicillin (500 mg tid) and metronidazole (400 mg tid). A strict supportive periodontal therapy regimen every three month in terms of supra- and subgingival debridement was rendered. The reported therapy has significantly improved periodontal health and arrested disease progression. Intraoral examination at the end of the observation period 3.5 years after non-surgical periodontal therapy showed generalized decreased pocket probing depths ranging from 1 to 6 mm, bleeding on probing lower 30%, and tooth mobility class I and II. Furthermore, microbiological analysis shows the absence of Porphyromonas gingivalis, Prevotella intermedia and Treponema denticola after therapy.
CONCLUSION: Adjunctive antibiotic treatment may alter the oral microbiome and thus, the inflammatory response of periodontal disease associated to plasminogen deficiency and diminishes the risk of pseudomembrane formation and progressive attachment loss. This case report indicates that patients with plasminogen deficiency may benefit from non-surgical periodontal treatment in combination with an adjunctive antibiotic therapy and a strict supportive periodontal therapy regimen.
A hallmark of several major neurological diseases is neuronal cell death. In addition to this primary pathology, secondary injury is seen in connected brain regions in which neurons not directly affected by the disease are denervated. These transneuronal effects on the network contribute considerably to the clinical symptoms. Since denervated neurons are viable, they are attractive targets for intervention. Therefore, we studied the role of Sphingosine-1-phosphate (S1P)-receptor signaling, the target of Fingolimod (FTY720), in denervation-induced dendritic atrophy. The entorhinal denervation in vitro model was used to assess dendritic changes of denervated mouse dentate granule cells. Live-cell microscopy of GFP-expressing granule cells in organotypic entorhino-hippocampal slice cultures was employed to follow individual dendritic segments for up to 6 weeks after deafferentation. A set of slice cultures was treated with FTY720 or the S1P-receptor (S1PR) antagonist VPC23019. Lesion-induced changes in S1P (mass spectrometry) and S1PR-mRNA levels (laser microdissection and qPCR) were determined. Denervation caused profound changes in dendritic stability. Dendritic elongation and retraction events were markedly increased, resulting in a net reduction of total dendritic length (TDL) during the first 2 weeks after denervation, followed by a gradual recovery in TDL. These changes were accompanied by an increase in S1P and S1PR1- and S1PR3-mRNA levels, and were not observed in slice cultures treated with FTY720 or VPC23019. We conclude that inhibition of S1PR signaling prevents dendritic destabilization and denervation-induced dendrite loss. These results suggest a novel neuroprotective effect for pharmaceuticals targeting neural S1PR pathways.
In common with several other autoimmune diseases, autoimmune Addison's disease (AAD) is thought to be caused by a combination of deleterious susceptibility polymorphisms in several genes, together with undefined environmental factors and stochastic events. To date, the strongest genomic association with AAD has been with alleles at the HLA locus, DR3-DQ2 and DR4. The contribution of other genetic variants has been inconsistent. We have studied the association of 16 single-nucleotide polymorphisms (SNPs) within the CD28-CTLA-4-ICOS genomic locus, in a cohort comprising 691 AAD patients of Norwegian and UK origin with matched controls. We have also performed a meta-analysis including 1002 patients from European countries. The G-allele of SNP rs231775 in CTLA-4 is associated with AAD in Norwegian patients (odds ratio (OR)=1.35 (confidence interval (CI) 1.10-1.66), P=0.004), but not in UK patients. The same allele is associated with AAD in the total European population (OR=1.37 (CI 1.13-1.66), P=0.002). A three-marker haplotype, comprising PROMOTER_1661, rs231726 and rs1896286 was found to be associated with AAD in the Norwegian cohort only (OR 2.43 (CI 1.68-3.51), P=0.00013). This study points to the CTLA-4 gene as a susceptibility locus for the development of AAD, and refines its mapping within the wider genomic locus.
A 79 year old female patient was admitted to our emergency department with a fracture of the right medial femoral neck six days after a fall on her right side and a cemented hemiprosthesis was implanted. Five days later, she developed a hemorrhagic shock and was diagnosed with a delayed splenic rupture and the spleen was resected. Histopathological examination showed a delayed rupture of an otherwise normal spleen without signs of an underlying pathology. The outcome was fatal: In the postoperative course she developed pneumonia, three weeks later she succumbed due to multiple organ failure.
Even careful reevaluation of the case did not provide any clues to expect an injury of the spleen according to trauma mechanism.
This case shows that delayed splenic rupture of a normal spleen may occur even after a low energy trauma. Injury of the spleen should therefore always be considered, even with an uncharacteristic anamnesis. Physical examination after trauma should therefore always include a careful clinical evaluation. The clinical threshold for a FAST examination should be low.
The coincidence of a femoral neck fracture and a splenic rupture after a low energy trauma has not been reported before.
Background The endogenous amino acid homoarginine predicts mortality in cerebro‐ and cardiovascular disease. The objective was to explore whether homoarginine is associated with atrial fibrillation (AF) and outcome in patients with acute chest pain.
Methods and Results One thousand six hundred forty‐nine patients with acute chest pain were consecutively enrolled in this study, of whom 589 were diagnosed acute coronary syndrome (ACS). On admission, plasma concentrations of homoarginine as well as brain natriuretic peptide (BNP), and high‐sensitivity assayed troponin I (hsTnI) were determined along with electrocardiography (ECG) variables. During a median follow‐up of 183 days, 60 major adverse cardiovascular events (MACEs; 3.8%), including all‐cause death, myocardial infarction, or stroke, were registered in the overall study population and 43 MACEs (7.5%) in the ACS subgroup. Adjusted multivariable Cox regression analyses revealed that an increase of 1 SD of plasma log‐transformed homoarginine (0.37) was associated with a hazard reduction of 26% (hazard ratio [HR], 0.74; 95% CI, 0.57–0.96) for incident MACE and likewise of 35% (HR, 0.65; 95% CI, 0.49–0.88) in ACS patients. In Kaplan–Meier survival curves, homoarginine was predictive for patients with high‐sensitivity assayed troponin I (hsTnI) above 27 ng/L (P<0.05). Last, homoarginine was inversely associated with QTc duration (P<0.001) and prevalent AF (OR, 0.83; 95% CI, 0.71–0.95).
Conclusion Low plasma homoarginine was identified as a risk marker for incident MACEs in patients with acute chest pain, in particular, in those with elevated hsTnI. Impaired homoarginine was associated with prevalent AF. Further studies are needed to investigate the link to AF and evaluate homoarginine as a therapeutic option for these patients.
BACKGROUND: We retrospectively evaluated the efficacy and toxicity of gross tumor volume (GTV) mean-dose-optimized and real-time motion-compensated robotic stereotactic body radiation therapy (SBRT) in the treatment of liver metastases.
METHODS: Between March 2011 and July 2015, 52 patients were treated with SBRT for a total of 91 liver metastases (one to four metastases per patient) with a median GTV volume of 12 cc (min 1 cc, max 372 cc). The optimization of mean GTV dose was prioritized during treatment planning at the potential cost of planning target volume (PTV) coverage reduction while adhering to safe normal tissue constraints. The delivered median GTV biological effective dose (BED10) was 142.1 Gy10 (range, 60.2 Gy10 -165.3 Gy10) and the prescribed PTV BED10 ranged from 40.6 Gy10 to 112.5 Gy10 (median, 86.1 Gy10). We analyzed local control (LC), progression-free interval (PFI), overall survival (OS), and toxicity.
RESULTS: Median follow-up was 17 months (range, 2-49 months). The 2-year actuarial LC, PFI, and OS rates were 82.1, 17.7, and 45.0 %, and the median PFI and OS were 9 and 23 months, respectively. In univariate analysis histology (p < 0.001), PTV prescription BED10 (HR 0.95, CI 0.91-0.98, p = 0.002) and GTV mean BED10 (HR 0.975, CI 0.954-0.996, p = 0.011) were predictive for LC. Multivariate analysis showed that only extrahepatic disease status at time of treatment was a significant factor (p = 0.033 and p = 0.009, respectively) for PFI and OS. Acute nausea or fatigue grade 1 was observed in 24.1 % of the patients and only 1 patient (1.9 %) had a side effect of grade ≥ 2.
CONCLUSIONS: Robotic real-time motion-compensated SBRT is a safe and effective treatment for one to four liver metastases. Reducing the PTV prescription dose and keeping a high mean GTV dose allowed the reduction of toxicity while maintaining a high local control probability for the treated lesions.
Cortical development extensively depends on sensory experience. Effects of congenital monaural and binaural deafness on cortical aural dominance and representation of binaural cues were investigated in the present study. We used an animal model that precisely mimics the clinical scenario of unilateral cochlear implantation in an individual with single-sided congenital deafness. Multiunit responses in cortical field A1 to cochlear implant stimulation were studied in normal-hearing cats, bilaterally congenitally deaf cats (CDCs), and unilaterally deaf cats (uCDCs). Binaural deafness reduced cortical responsiveness and decreased response thresholds and dynamic range. In contrast to CDCs, in uCDCs, cortical responsiveness was not reduced, but hemispheric-specific reorganization of aural dominance and binaural interactions were observed. Deafness led to a substantial drop in binaural facilitation in CDCs and uCDCs, demonstrating the inevitable role of experience for a binaural benefit. Sensitivity to interaural time differences was more reduced in uCDCs than in CDCs, particularly at the hemisphere ipsilateral to the hearing ear. Compared with binaural deafness, unilateral hearing prevented nonspecific reduction in cortical responsiveness, but extensively reorganized aural dominance and binaural responses. The deaf ear remained coupled with the cortex in uCDCs, demonstrating a significant difference to deprivation amblyopia in the visual system.
Novel insights into the regulation of cyclooxygenase-2 expression by platelet-cancer cell cross-talk
(2015)
Platelets are activated by the interaction with cancer cells and release enhanced levels of lipid mediators [such as thromboxane (TX)A2 and prostaglandin (PG)E2, generated from arachidonic acid (AA) by the activity of cyclooxygenase (COX)-1], granule content, including ADP and growth factors, chemokines, proteases and Wnt proteins. Moreover, activated platelets shed different vesicles, such as microparticles (MPs) and exosomes (rich in genetic material such as mRNAs and miRNAs). These platelet-derived products induce several phenotypic changes in cancer cells which confer high metastatic capacity. A central event involves an aberrant expression of COX-2 which influences cell-cycle progression and contribute to the acquisition of a cell migratory phenotype through the induction of epithelial mesenchymal transition genes and down-regulation of E-cadherin expression. The identification of novel molecular determinants involved in the cross-talk between platelets and cancer cells has led to identify novel targets for anti-cancer drug development.
The wide range of immunosuppressive therapies and protocols permits tailored planning of the initial regimen according to the immunological risk status of individual patients. Pre-transplant risk assessment can include many factors, but there is no clear consensus on which parameters to take into account, and their relative importance. In general younger patients are known to be at higher risk for acute rejection, compounded by higher rates of non-adherence in adolescents. Donor age and recipient gender do not appear to exert a meaningful effect on risk of rejection per se, but black recipient ethnicity remains a well-established risk factor even under modern immunosuppression regimens. Little difference in risk is now observed between deceased- and living-donor recipients. Immunological risk assessment has developed substantially in recent years. Cross-match testing with cytotoxic analysis has long been supplemented by flow cytometry, but development of solid-phase single-bead antigen testing of solubilized human leukocyte antigens (HLA) to detect donor-specific antibodies (DSA) permits a far more nuanced stratification of immunological risk status, including the different classes and intensities of HLA antibodies Class I and/or II, including HLA-DSA. Immunologic risk evaluation is now often based on a combination of these tests, but other assessments are becoming more widely introduced, such as measurement of non-HLA antibodies against angiotensin type 1 (AT1) receptors or T-cell ELISPOT assay of alloantigen-specific donor. Targeted densensitization protocols can improve immunological risk, notably for DSA-positive patients with negative cytotoxicity and flow cross-match. HLA mismatch remains an important and undisputed risk factor for rejection. Delayed graft function also increases the risk of subsequent acute rejection, and the early regimen can be modified in such cases. Overall, there is a shift towards planning the immunosuppressive regimen based on pre-transplant immunology testing although certain conventional risk factors retain their importance.
Background: It is occasionally difficult to distinguish the stellate reticulum-like region of ameloblastoma from the fibrous connective tissue stroma. This difficulty is further pronounced in the plexiform variant of ameloblastoma that has very sparse fibrous connective tissue.
Aim: To test the utility of Azan trichrome stain in marking tumour regions and the peri-tumour environment of ameloblastoma.
Materials and Methods: Sections were prepared for 18 formalin fixed paraffin-embedded blocks of ameloblastoma cases and stained with Azan trichrome stain according to the manufacturer's specification.
Results and Conclusions: The tumour areas were stained mostly brown, with the ameloblasts mainly marked as deep brown while the stellate reticulum-like region was light brown. The structures in the peri-tumour region were marked with different shades of blue. Azan trichrome staining was able to distinguish between the fibrous connective tissue and the stellate reticulum-like areas in 100% of the cases.
Introduction: ameloblastoma is a slow growing, painless odontogenic swelling which can attain sizes that result in severe deformities of the craniofacial complex. It is the most commonly encountered odontogenic tumor in Nigeria. Surgical intervention is currently the method of treatment; however identification of altered molecular pathways may inform chemotherapeutic potential. The Protein Patched homolog 1 (PTCH-1) is overexpressed in ameloblastoma. Also, mutation in the MDM2 gene can reduce the tumor suppressor function of p53 and promote ameloblastoma growth. No study however has characterized the molecular profile of African cases of ameloblastoma with a view to developing chemotherapeutic alternatives. The objective was to characterize the PTCH-1 genetic profile of Ameloblastoma in Nigerian patients as a first step in investigating its potential for chemotherapeutic intervention.
Methods: twenty-eight FFPE blocks of ameloblastoma cases from Nigerian patients were prepared for antibody processing to PTCH-1 (Polyclonal Anti-PTCH antibody ab39266) and MDM2 (Monoclonal Anti-MDM2 antibody (2A10) ab16895). Cytoplasmic brown staining was considered as positive for PTCH while nuclear staining was positive for MDM2.
Results: moderate and strong expressions for PTCH in ameloblast and stellate reticulum were 78.6% and 60.7% respectively. Only 3 (10.7%) cases expressed MDM2.
Conclusion: the importance of our study is that it supports, in theory, anti-PTCH/SHH chemotherapeutics for Nigerian ameloblastoma cases and also infers the possible additional use of anti-p53 agents.
Protein turnover and quality control by the proteasome is of paramount importance for cell homeostasis. Dysfunction of the proteasome is associated with aging processes and human diseases such as neurodegeneration, cardiomyopathy, and cancer. The regulation, i.e. activation and inhibition of this fundamentally important protein degradation system, is still widely unexplored. We demonstrate here that the evolutionarily highly conserved type II triple-A ATPase VCP and the proteasome inhibitor PSMF1/PI31 interact directly, and antagonistically regulate proteasomal activity. Our data provide novel insights into the regulation of proteasomal activity.
Polo-like kinase 1 regulates the stability of the mitotic centromere-associated kinesin in mitosis
(2014)
Proper bi-orientation of chromosomes is critical for the accurate segregation of chromosomes in mitosis. A key regulator of this process is MCAK, the mitotic centromere-associated kinesin. During mitosis the activity and localization of MCAK are regulated by mitotic key kinases including Plk1 and Aurora B. We show here that S621 in the MCAK’s C-terminal domain is the major phosphorylation site for Plk1. This phosphorylation regulates MCAK’s stability and facilitates its recognition by the ubiquitin/proteasome dependent APC/CCdc20 pathway leading to its D-box dependent degradation in mitosis. While phosphorylation of S621 does not directly affect its microtubule depolymerising activity, loss of Plk1 phosphorylation on S621 indirectly enhances its depolymerization activity in vivo by stabilizing MCAK, leading to an increased level of protein. Interfering with phosphorylation at S621 causes spindle formation defects and chromosome misalignments. Therefore, this study suggests a new mechanism by which Plk1 regulates MCAK: by regulating its degradation and hence controlling its turnover in mitosis.
MiR144/451 expression is repressed by RUNX1 during megakaryopoiesis and disturbed by RUNX1/ETO
(2016)
Abstract: A network of lineage-specific transcription factors and microRNAs tightly regulates differentiation of hematopoietic stem cells along the distinct lineages. Deregulation of this regulatory network contributes to impaired lineage fidelity and leukemogenesis. We found that the hematopoietic master regulator RUNX1 controls the expression of certain microRNAs, of importance during erythroid/megakaryocytic differentiation. In particular, we show that the erythorid miR144/451 cluster is epigenetically repressed by RUNX1 during megakaryopoiesis. Furthermore, the leukemogenic RUNX1/ETO fusion protein transcriptionally represses the miR144/451 pre-microRNA. Thus RUNX1/ETO contributes to increased expression of miR451 target genes and interferes with normal gene expression during differentiation. Furthermore, we observed that inhibition of RUNX1/ETO in Kasumi1 cells and in RUNX1/ETO positive primary acute myeloid leukemia patient samples leads to up-regulation of miR144/451. RUNX1 thus emerges as a key regulator of a microRNA network, driving differentiation at the megakaryocytic/erythroid branching point. The network is disturbed by the leukemogenic RUNX1/ETO fusion product.
Author Summary: The regulatory network between transcription factors, epigenetic cofactors and microRNAs is decisive for normal hematopoiesis. The transcription factor RUNX1 is important for the establishment of a megakaryocytic gene expression program and the concomitant repression of erythroid genes during megakaryocytic differentiation. Gene regulation by RUNX1 is frequently disturbed by mutations and chromosomal translocations, such as the t(8;21) translocation, which gives rise to the leukemogenic RUNX1/ETO fusion protein. We found that RUNX1 regulates microRNAs, which are of importance at the megakaryocytic/erythroid branching point. Specifically, RUNX1 down-regulates expression of the microRNA cluster miR144/451 during megakaryocytic differentiation by changing the epigenetic histone modification pattern at the locus. We could show that miR451, one of the micorRNAs of the miR144/451 cluster, supports erythroid differentiation. We found that expression of miR451 is repressed by the RUNX1/ETO fusion protein, resulting in up regulation of miR451 target genes. Our data support the notion that RUNX1 suppresses the erythroid gene expression program including the erythroid microRNA miR451 and that the RUNX1/ETO fusion protein interferes with normal gene regulation by RUNX1.
Aims: To investigate real-world clinical and patient-related variables associated with initiating GLP-1 receptor agonist (GLP-1RA) treatment relative to initiation of other glucose-lowering therapies in type 2 diabetes (T2D) patients of primary care in Germany.
Methods: Data for 938 T2D patients who started therapy with a GLP-1RA within 823 practices of primary care throughout Germany were retrospectively analyzed (Disease Analyser: 01/2011–03/2014). 5,197 T2D patients who initiated other non-GLP-1RA antidiabetic therapies were selected as controls. Multivariate logistic regression analyses were applied to identify factors associated with GLP-1RA initiation in primary care.
Results: Mean age (SD) of GLP-1RA users was 57.8 (11.8) years (males: 55.5%) and the average BMI was 36.1 (6.7) kg/m2. 22.8% were in diabetologist care and 12.0% had private health insurance. In multivariate regression, choice of GLP-1RA therapy instead of a different glucose-lowering drug class was associated with obesity (odds ratio: 1.68; 95% CI: 1.34–2.10), private health insurance (2.42; 1.89–3.09), younger age (0.94; 0.93–0.95 per year), male sex (0.85; 0.73–0.99), diabetologist care (2.11; 1.73–2.57), and geographic practice location (East vs. West-Germany; 1.25; 1.05–1.49). Among co-medication, angiotensin II antagonists (increased) and non-steroidal antirheumatic agents (decreased) were related to GLP-1RA prescriptions (both p<0.001).
Conclusions: Consistent with German guidelines, GLP-1RA is mainly prescribed preferentially in T2D patients who are obese. GLP-1RA drugs were more frequently used than other options in privately health insured patients and in patients seeing a diabetologist.
Background: We conducted a phase I study with a granulocyte macrophage colony stimulating factor (GMCSF)-expressing oncolytic adenovirus, ONCOS-102, in patients with solid tumors refractory to available treatments. The objectives of the study were to determine the optimal dose for further use and to assess the safety, tolerability and adverse event (AE) profile of ONCOS-102. Further, the response rate and overall survival were evaluated as well as preliminary evidence of disease control. As an exploratory endpoint, the effect of ONCOS 102 on biological correlates was examined.
Methods: The study was conducted using a classic 3 + 3 dose escalation study design involving 12 patients. Patients were repeatedly treated intratumorally with ONCOS-102 plus daily low-dose oral cyclophosphamide (CPO). Tumor response was evaluated with diagnostic positron emission tomography (PET) and computed tomography (CT). Tumor biopsies were collected at baseline and after treatment initiation for analysis of immunological correlates. Peripheral blood mononuclear cells (PBMCs) were collected at baseline and during the study to assess antigen specificity of CD8+ T cells by interferon gamma (IFNγ) enzyme linked immunospot assay (ELISPOT).
Results: No dose limiting toxicity (DLT) or maximum tolerated dose (MTD) was identified for ONCOS-102. Four out of ten (40 %) evaluable patients had disease control based on PET/CT scan at 3 months and median overall survival was 9.3 months. A short-term increase in systemic pro-inflammatory cytokines and a prominent infiltration of TILs to tumors was seen post-treatment in 11 out of 12 patients. Two patients showed marked infiltration of CD8+ T cells to tumors and concomitant systemic induction of tumor-specific CD8+ T cells. Interestingly, high expression levels of genes associated with activated TH1 cells and TH1 type immune profile were observed in the post-treatment biopsies of these two patients.
Conclusions: ONCOS-102 is safe and well tolerated at the tested doses. All three examined doses may be used in further development. There was evidence of antitumor immunity and signals of clinical efficacy. Importantly, treatment resulted in infiltration of CD8+ T cells to tumors and up-regulation of PD-L1, highlighting the potential of ONCOS-102 as an immunosensitizing agent for combinatory therapies with checkpoint inhibitors.
Trial registration: NCT01598129. Registered 19/04/2012
Objective: To investigate the impact of HPV status in patients with locally advanced head and neck squamous cell carcinoma (HNSCC), who received surgery and cisplatin-based postoperative radiochemotherapy.
Materials and methods: For 221 patients with locally advanced squamous cell carcinoma of the hypopharynx, oropharynx or oral cavity treated at the 8 partner sites of the German Cancer Consortium, the impact of HPV DNA, p16 overexpression and p53 expression on outcome were retrospectively analysed. The primary endpoint was loco-regional tumour control; secondary endpoints were distant metastases and overall survival.
Results: In the total patient population, univariate analyses revealed a significant impact of HPV16 DNA positivity, p16 overexpression, p53 positivity and tumour site on loco-regional tumour control. Multivariate analysis stratified for tumour site showed that positive HPV 16 DNA status correlated with loco-regional tumour control in patients with oropharyngeal carcinoma (p = 0.02) but not in the oral cavity carcinoma group. Multivariate evaluation of the secondary endpoints in the total population revealed a significant association of HPV16 DNA positivity with overall survival (p < 0.01) but not with distant metastases.
Conclusions: HPV16 DNA status appears to be a strong prognosticator of loco-regional tumour control after postoperative cisplatin-based radiochemotherapy of locally advanced oropharyngeal carcinoma and is now being explored in a prospective validation trial.
Late stage cancer is often associated with reduced immune recognition and a highly immunosuppressive tumor microenvironment. The presence of tumor infiltrating lymphocytes (TILs) and specific gene-signatures prior to treatment are linked to good prognosis, while the opposite is true for extensive immunosuppression. The use of adenoviruses as cancer vaccines is a form of active immunotherapy to initialise a tumor-specific immune response that targets the patient’s unique tumor antigen repertoire. We report a case of a 68-year-old male with asbestos-related malignant pleural mesothelioma who was treated in a Phase I study with a granulocyte-macrophage colony‑stimulating factor (GM-CSF)-expressing oncolytic adenovirus, Ad5/3-D24-GMCSF (ONCOS-102). The treatment resulted in prominent infiltration of CD8C lymphocytes to tumor, marked induction of systemic antitumor CD8C T-cells and induction of Th1- type polarization in the tumor. These results indicate that ONCOS-102 treatment sensitizes tumors to other immunotherapies by inducing a T-cell positive phenotype to an initially T-cell negative tumor.
Bbackground: We aimed to investigate the prognostic value of tumour-infiltrating lymphocytes’ (TILs) expression in pretreatment specimens from patients with head and neck squamous cell carcinoma (HNSCC) treated with definitive chemoradiotherapy (CRT).
Methods: The prevalence of CD3+, CD8+, CD4+ and FOXP3+ TILs was assessed using immunohistochemistry in tumour tissue obtained from 101 patients before CRT and was correlated with clinicopathological characteristics as well as local failure-free- (LFFS), distant metastases free- (DMFS), progression-free (PFS) and overall survival (OS). Survival curves were measured using the Kaplan–Meier method, and differences in survival between the groups were estimated using the log-rank test. Prognostic effects of TIL subset density were determined using the Cox regression analysis.
Results: With a mean follow-up of 25 months (range, 2.3–63 months), OS at 2 years was 57.4% for the entire cohort. Patients with high immunohistochemical CD3 and CD8 expression had significantly increased OS (P=0.024 and P=0.028), PFS (P=0.044 and P=0.047) and DMFS (P=0.021 and P=0.026) but not LFFS (P=0.90 and P=0.104) in multivariate analysis that included predictive clinicopathologic factors, such as age, sex, T-stage, N-stage, tumour grading and localisation. Neither CD4 nor FOXP3 expression showed significance for the clinical outcome. The lower N-stage was associated with improved OS in the multivariate analysis (P=0.049).
Conclusion: The positive correlation between a high number of infiltrating CD3+ and CD8+ cells and clinical outcome indicates that TILs may have a beneficial role in HNSCC patients and may serve as a biomarker to identify patients likely to benefit from definitive CRT.
The translation inhibitor and tumor suppressor Pdcd4 was reported to be lost in various tumors and put forward as prognostic marker in tumorigenesis. Decreased Pdcd4 protein stability due to PI3K-mTOR-p70S6K1 dependent phosphorylation of Pdcd4 followed by β-TrCP1-mediated ubiquitination, and proteasomal destruction of the protein was characterized as a major mechanism contributing to the loss of Pdcd4 expression in tumors. In an attempt to identify stabilizers of Pdcd4, we used a luciferase-based high-throughput compatible cellular assay to monitor phosphorylation-dependent proteasomal degradation of Pdcd4 in response to mitogen stimulation. Following a screen of approximately 2000 compounds, we identified 1,2-bis(4-chlorophenyl)disulfide as a novel Pdcd4 stabilizer. To determine an initial structure-activity relationship, we used 3 additional compounds, synthesized according to previous reports, and 2 commercially available compounds for further testing, in which either the linker between the aryls was modified (compounds 2–4) or the chlorine residues were replaced by groups with different electronic properties (compounds 5 and 6). We observed that those compounds with alterations in the sulfide linker completely lost the Pdcd4 stabilizing potential. In contrast, modifications in the chlorine residues showed only minor effects on the Pdcd4 stabilizing activity. A reporter with a mutated phospho-degron verified the specificity of the compounds for stabilizing the Pdcd4 reporter. Interestingly, the active diaryl disulfides inhibited proliferation and viability at concentrations where they stabilized Pdcd4, suggesting that Pdcd4 stabilization might contribute to the anti-proliferative properties. Finally, computational modelling indicated that the flexibility of the disulfide linker might be necessary to exert the biological functions of the compounds, as the inactive compound appeared to be energetically more restricted.
Even though extensively investigated, the nature of working memory (WM) deficits in patients with schizophrenia (PSZ) is not yet fully understood. In particular, the contribution of different WM sub-processes to the severe WM deficit observed in PSZ is a matter of debate. So far, most research has focused on impaired WM maintenance. By analyzing different types of errors in a spatial delayed response task (DRT), we have recently demonstrated that incorrect yet confident responses (which we labeled as false memory errors) rather than incorrect/not-confident responses reflect failures of WM encoding, which was also impaired in PSZ. In the present study, we provide further evidence for a functional dissociation between confident and not-confident errors by manipulating the demands on WM maintenance, i.e., the length over which information has to be maintained in WM. Furthermore, we investigate whether these functionally distinguishable WM processes are impaired in PSZ. Twenty-four PSZ and 24 demographically matched healthy controls (HC) performed a spatial DRT in which the length of the delay period was varied between 1, 2, 4, and 6 s. In each trial, participants also rated their level of response confidence. Across both groups, longer delays led to increased rates of incorrect/not-confident responses, while incorrect/confident responses were not affected by delay length. This functional dissociation provides additional support for our proposal that false memory errors (i.e., confident errors) reflect problems at the level of WM encoding, while not-confident errors reflect failures of WM maintenance. Schizophrenic patients showed increased numbers of both confident and not-confident errors, suggesting that both sub-processes of WM—encoding and maintenance—are impaired in schizophrenia. Combined with the delay length-dependent functional dissociation, we propose that these impairments in schizophrenic patients are functionally distinguishable.
Abstract:
Snakebite is an important medical emergency in rural Nepal. Correct identification of the biting species is crucial for clinicians to choose appropriate treatment and anticipate complications. This is particularly important for neurotoxic envenoming which, depending on the snake species involved, may not respond to available antivenoms. Adequate species identification tools are lacking. This study used a combination of morphological and molecular approaches (PCR-aided DNA sequencing from swabs of bite sites) to determine the contribution of venomous and non-venomous species to the snakebite burden in southern Nepal. Out of 749 patients admitted with a history of snakebite to one of three study centres, the biting species could be identified in 194 (25.9%). Out of these, 87 had been bitten by a venomous snake, most commonly the Indian spectacled cobra (Naja naja; n = 42) and the common krait (Bungarus caeruleus; n = 22). When both morphological identification and PCR/sequencing results were available, a 100% agreement was noted. The probability of a positive PCR result was significantly lower among patients who had used inadequate “first aid” measures (e.g. tourniquets or local application of remedies). This study is the first to report the use of forensic genetics methods for snake species identification in a prospective clinical study. If high diagnostic accuracy is confirmed in larger cohorts, this method will be a very useful reference diagnostic tool for epidemiological investigations and clinical studies.
Author Summary:
Snakebite is an important medical problem in sub-tropical and tropical regions, including Nepal where tens of thousands of people are bitten every year. Snakebite can result in life-threatening envenoming, and correct identification of the biting species is crucial for care providers to choose appropriate treatment and anticipate complications. This paper explores a number of methods, including molecular techniques, to assist care providers in identifying the species responsible for bites in rural Nepal. Out of 749 patients with a history of snakebite, the biting species could be identified in 194 (25.9%). Out of these, 87 had been bitten by a venomous snake, most commonly cobras (n = 42) and kraits (n = 22). This study is the first to report the use of molecular techniques for snake species identification. The diagnostic accuracy of this method appears high but needs to be confirmed in larger studies.
Purpose: To evaluate the efficacy of the virtual reality training simulator Eyesi to prepare surgeons for performing pars plana vitrectomies and its potential to predict the surgeons’ performance.
Methods: In a preparation phase, four participating vitreoretinal surgeons performed repeated simulator training with predefined tasks. If a surgeon was assigned to perform a vitrectomy for the management of complex retinal detachment after a surgical break of at least 60 hours it was randomly decided whether a warmup training on the simulator was required (n = 9) or not (n = 12). Performance at the simulator was measured using the built-in scoring metrics. The surgical performance was determined by two blinded observers who analyzed the video-recorded interventions. One of them repeated the analysis to check for intra-observer consistency. The surgical performance of the interventions with and without simulator training was compared. In addition, for the surgeries with simulator training, the simulator performance was compared to the performance in the operating room.
Results: Comparing each surgeon’s performance with and without warmup trainingshowed a significant effect of warmup training onto the final outcome in the operating room. For the surgeries that were preceeded by the warmup procedure, the performance at the simulator was compared with the operating room performance. We found that there is a significant relation. The governing factor of low scores in the simulator were iatrogenic retinal holes, bleedings and lens damage. Surgeons who caused minor damage in the simulation also performed well in the operating room.
Conclusions: Despite the large variation of conditions, the effect of a warmup training as well as a relation between the performance at the simulator and in the operating room was found with statistical significance. Simulator training is able to serve as a warmup to increase the average performance.
Background and Objectives: Tick-borne encephalitis (TBE) still represents a considerable medical and health economic problem in Europe and entails a potential threat to travellers. The aim of this study was to characterise the conditions of severe TBE by precisely recording its clinical variants, the related neuroimaging features, and the variant-specific long-term outcome and by identifying predictors for severe courses.
Methods: A cohort of 111 TBE patients (median age 51, range 17–75 years; 42% females) was analysed prospectively. Data were acquired from the department of neurology, University Hospital Heidelberg, and the infectious diseases registry of the Robert-Koch institute Berlin. Neurological status was ascertained by protocol at admission and discharge and the degree of disability was scored using the modified RANKIN Scale (mRS; clinical score addressing neurological disability, range from 0, healthy to 6, dead) at admission and at follow-up. Follow-up examination was conducted by means of a telephone interview. To identify independent predictors for severe TBE and functional outcome, modelled logistic regression was performed. MRI changes were correlated with infection variants. To assess alpha-motor neuron injury patterns, we used high resolution magnetic resonance neurography (hrMRN). Analyses were performed at the Department of Neurology, University Hospital, University of Heidelberg from April 2004 through September 2014
Results: Acute course: 3.6% of patients died during the acute infection. All patients with a lethal course suffered from meningoencephaloradiculitis (MER, 14.4% of the cohort), which is associated with a significantly higher risk of requiring intensive care (p = 0.004) and mechanical ventilation (p<0.001) than menigoencephalitis (ME, 27.9% of the cohort). At admission, both MER and ME groups were severely affected, with the MER group having a statistically higher mRS score (median of 5 in the MER groups versus 4 in the ME group; p<0.001). Long-term outcome: outcome for MER was considerably worse (median mRS = 4) than for ME (mRS = 1, p<0.0001) and meningitis (mRS = 0, 57.7% of the cohort). Risk factors: advanced age (p<0.001) and male gender (p = 0.043) are independent risk factors for a severe infection course. Furthermore, we identified pre-existing diabetes mellitus (p = 0.024) as an independent risk factor for MER. In MER, alpha-motor neuron injury accounts for the poor prognosis confirmed by hrMRN.
Conclusion and Relevance: These data provide critical information for neurologists and other health professionals to use in evaluating TBEV patients who live in or travel to endemic areas. This information can be used to classify clinical presentation and estimate infection-associated complications and individual prognosis. Furthermore, the risk for severe, disabling infections in older patients should prompt general practitioners to recommend and encourage vaccination to those patients living in or travelling to endemic areas.
We previously reported that aberrant HH pathway activation confers a poor prognosis in rhabdomyosarcoma (RMS). Searching for new treatment strategies we therefore targeted HH signaling. Here, we identify a novel synthetic lethality of concomitant inhibition of HH and PI3K/AKT/mTOR pathways in RMS by GLI1/2 inhibitor GANT61 and PI3K/mTOR inhibitor PI103. Synergistic drug interaction is confirmed by calculation of combination index (CI < 0.2). Similarly, genetic silencing of GLI1/2 significantly increases PI103-induced apoptosis. GANT61 and PI103 also synergize to induce apoptosis in cultured primary RMS cells emphasizing the clinical relevance of this combination. Importantly, GANT61/PI103 cotreatment suppresses clonogenic survival, three-dimensional sphere formation and tumor growth in an in vivo model of RMS. Mechanistic studies reveal that GANT61 and PI103 cooperate to trigger caspase-dependent apoptosis via the mitochondrial pathway, as demonstrated by several lines of evidence. First, GANT61/PI103 cotreatment increases mRNA and protein expression of NOXA and BMF, which is required for apoptosis, since knockdown of NOXA or BMF significantly reduces GANT61/PI103-induced apoptosis. Second, GANT61/PI103 cotreatment triggers BAK/BAX activation, which contributes to GANT61/PI103-mediated apoptosis, since knockdown of BAK provides protection. Third, ectopic expression of BCL-2 or non-degradable phospho-mutant MCL-1 significantly rescue GANT61/PI103-triggered apoptosis. Fourth, GANT61/PI103 cotreatment initiate activation of the caspase cascade via apoptosome-mediated cleavage of the initiator caspase-9, as indicated by changes in the cleavage pattern of caspases (e.g. accumulation of the caspase-9 p35 cleavage fragment) upon addition of the caspase inhibitor zVAD.fmk. Thus, combined GLI1/2 and PI3K/mTOR inhibition represents a promising novel approach for synergistic apoptosis induction and tumor growth reduction with implications for new treatment strategies in RMS.
After the mass-vaccination campaign during the influenza A (H1N1) 2009 pandemic, a significant increase in narcolepsy incidence was observed initially in Scandinavia, later in other European countries and recently also in Canada. Narcolepsy is a sleep disease caused by the loss of hypocretin-producing cells in the hypothalamus. Almost all narcolepsy patients carry the HLA-DQB1*0602 allele, giving a link to an autoimmune-mediated process.
Most of the observed narcolepsy cases were correlated to the vaccination with Pandemrix, the most frequently used vaccine in the EU, and a slight connection to Arepanrix was also detected, which was distributed in Canada. Both vaccines were adjuvanted with AS03, suggesting a possible link between AS03 and narcolepsy. No narcolepsy cases were detected with MF59-adjuvanted or non-adjuvanted influenza vaccines. Recent studies reported differences between Pandemrix and Arepanrix and suggested the vaccine rather than the adjuvant as a suspect for narcolepsy development following vaccination. In addition, in China an increase of narcolepsy cases was reported to occur in absence of vaccination. Possible factors and potential additive effects that may have triggered narcolepsy after the pandemic vaccination are being reviewed in this paper.
Biomedical data obtained during cell experiments, laboratory animal research, or human studies often display a complex distribution. Statistical identification of subgroups in research data poses an analytical challenge. Here were introduce an interactive R-based bioinformatics tool, called “AdaptGauss”. It enables a valid identification of a biologically-meaningful multimodal structure in the data by fitting a Gaussian mixture model (GMM) to the data. The interface allows a supervised selection of the number of subgroups. This enables the expectation maximization (EM) algorithm to adapt more complex GMM than usually observed with a noninteractive approach. Interactively fitting a GMM to heat pain threshold data acquired from human volunteers revealed a distribution pattern with four Gaussian modes located at temperatures of 32.3, 37.2, 41.4, and 45.4 °C. Noninteractive fitting was unable to identify a meaningful data structure. Obtained results are compatible with known activity temperatures of different TRP ion channels suggesting the mechanistic contribution of different heat sensors to the perception of thermal pain. Thus, sophisticated analysis of the modal structure of biomedical data provides a basis for the mechanistic interpretation of the observations. As it may reflect the involvement of different TRP thermosensory ion channels, the analysis provides a starting point for hypothesis-driven laboratory experiments.
It is now accepted that heart failure (HF) is a complex multifunctional disease rather than simply a hemodynamic dysfunction. Despite its complexity, stressed cardiomyocytes often follow conserved patterns of structural remodelling in order to adapt, survive, and regenerate. When cardiac adaptations cannot cope with mechanical, ischemic, and metabolic loads efficiently or become chronically activated, as, for example, after infection, then the ongoing structural remodelling and dedifferentiation often lead to compromised pump function and patient death. It is, therefore, of major importance to understand key events in the progression from a compensatory left ventricular (LV) systolic dysfunction to a decompensatory LV systolic dysfunction and HF. To achieve this, various animal models in combination with an “omics” toolbox can be used. These approaches will ultimately lead to the identification of an arsenal of biomarkers and therapeutic targets which have the potential to shape the medicine of the future.
The Sonic Hedgehog (SHH) pathway plays a central role in the developing mammalian CNS. In our study, we aimed to investigate the spatiotemporal SHH pathway expression pattern in human fetal brains. We analyzed 22 normal fetal brains for Shh, Patched, Smoothened, and Gli1-3 expression by immunohistochemistry. In the telencephalon, strongest expression of Shh, Smoothened, and Gli2 was found in the cortical plate (CP) and ventricular zone. Patched was strongly upregulated in the ventricular zone and Gli1 in the CP. In the cerebellum, SHH pathway members were strongly expressed in the external granular layer (EGL). SHH pathway members significantly decreased over time in the ventricular and subventricular zone and in the cerebellar EGL, while increasing levels were found in more superficial telencephalic layers. Our findings show that SHH pathway members are strongly expressed in areas important for proliferation and differentiation and indicate a temporal expression gradient in telencephalic and cerebellar layers probably due to decreased proliferation of progenitor cells and increased differentiation. Our data about the spatiotemporal expression of SHH pathway members in the developing human brain serves as a base for the understanding of both normal and pathological CNS development.
Psoriatic arthritis (PsA) is a chronic inflammatory arthritis associated with psoriasis and, despite the larger estimated heritability for PsA, the majority of genetic susceptibility loci identified to date are shared with psoriasis. Here, we present results from a case-control association study on 1,962 PsA patients and 8,923 controls using the Immunochip genotyping array. We identify eight loci passing genome-wide significance, secondary independent effects at three loci and a distinct PsA-specific variant at the IL23R locus. We report two novel loci and evidence of a novel PsA-specific association at chromosome 5q31. Imputation of classical HLA alleles, amino acids and SNPs across the MHC region highlights three independent associations to class I genes. Finally, we find an enrichment of associated variants to markers of open chromatin in CD8(+) memory primary T cells. This study identifies key insights into the genetics of PsA that could begin to explain fundamental differences between psoriasis and PsA.
Objective: To evaluate two ultrafast cone-beam CT (UF-CBCT) imaging protocols with different acquisition and injection parameters regarding image quality and required contrast media during image-guided hepatic transarterial chemoembolization (TACE).
Methods: In 80 patients (male: 46, female: 34; mean age: 56.8 years; range: 33–83) UF-CBCT was performed during TACE for intraprocedural guidance. Imaging was performed using two ultrafast CBCT acquisition protocols with different acquisition and injection parameters (imaging protocol 1: acquisition time 2.54 s, and contrast 6 mL with 3 s delay; imaging protocol 2: acquisition time 2.72 s, and contrast 7 mL with 6 s delay). Image evaluation was performed with both qualitative and quantitative methods. Contrast injection volume and dose parameters were compared using values from the literature.
Results: Imaging protocol 2 provided significantly better (P < 0.05) image quality than protocol 1 at the cost of slightly higher contrast load and patient dose. Imaging protocol 1 provided good contrast perfusion but it mostly failed to delineate the tumors (P < 0.05). On the contrary, imaging protocol 2 showed excellent enhancement of hepatic parenchyma, tumor, and feeding vessels.
Conclusion: Tumor delineation, visualization of hepatic parenchyma, and feeding vessels are clearly possible using imaging protocol 2 with ultrafast CBCT imaging. A reduction of required contrast volume and patient dose were achieved due to the ultrafast CBCT imaging.
In vivo long-term monitoring of circulating tumor cells fluctuation during medical interventions
(2015)
The goal of this research was to study the long-term impact of medical interventions on circulating tumor cell (CTC) dynamics. We have explored whether tumor compression, punch biopsy or tumor resection cause dissemination of CTCs into peripheral blood circulation using in vivo fluorescent flow cytometry and breast cancer-bearing mouse model inoculated with MDA-MB-231-Luc2-GFP cells in the mammary gland. Two weeks after tumor inoculation, three groups of mice were the subject of the following interventions: (1) tumor compression for 15 minutes using 400 g weight to approximate the pressure during mammography; (2) punch biopsy; or (3) surgery. The CTC dynamics were determined before, during and six weeks after these interventions. An additional group of tumor-bearing mice was used as control and did not receive an intervention. The CTC dynamics in all mice were monitored weekly for eight weeks after tumor inoculation. We determined that tumor compression did not significantly affect CTC dynamics, either during the procedure itself (P = 0.28), or during the 6-week follow-up. In the punch biopsy group, we observed a significant increase in CTC immediately after the biopsy (P = 0.02), and the rate stayed elevated up to six weeks after the procedure in comparison to the tumor control group. The CTCs in the group of mice that received a tumor resection disappeared immediately after the surgery (P = 0.03). However, CTC recurrence in small numbers was detected during six weeks after the surgery. In the future, to prevent these side effects of medical interventions, the defined dynamics of intervention-induced CTCs may be used as a basis for initiation of aggressive anti-CTC therapy at time-points of increasing CTC number.
At the sensor level many aspects, such as spectral power, functional and effective connectivity as well as relative-power-ratio ratio (RPR) and spatial resolution have been comprehensively investigated through both electroencephalography (EEG) and magnetoencephalography (MEG). Despite this, differences between both modalities have not yet been systematically studied by direct comparison. It remains an open question as to whether the integration of EEG and MEG data would improve the information obtained from the above mentioned parameters. Here, EEG (64-channel system) and MEG (275 sensor system) were recorded simultaneously in conditions with eyes open (EO) and eyes closed (EC) in 29 healthy adults. Spectral power, functional and effective connectivity, RPR, and spatial resolution were analyzed at five different frequency bands (delta, theta, alpha, beta and gamma). Networks of functional and effective connectivity were described using a spatial filter approach called the dynamic imaging of coherent sources (DICS) followed by the renormalized partial directed coherence (RPDC). Absolute mean power at the sensor level was significantly higher in EEG than in MEG data in both EO and EC conditions. At the source level, there was a trend towards a better performance of the combined EEG+MEG analysis compared with separate EEG or MEG analyses for the source mean power, functional correlation, effective connectivity for both EO and EC. The network of coherent sources and the spatial resolution were similar for both the EEG and MEG data if they were analyzed separately. Results indicate that the combined approach has several advantages over the separate analyses of both EEG and MEG. Moreover, by a direct comparison of EEG and MEG, EEG was characterized by significantly higher values in all measured parameters in both sensor and source level. All the above conclusions are specific to the resting state task and the specific analysis used in this study to have general conclusion multi-center studies would be helpful.
Adult neurogenesis is frequently studied in the mouse hippocampus. We examined the morphological development of adult-born, immature granule cells in the suprapyramidal blade of the septal dentate gyrus over the period of 7–77 days after mitosis with BrdU-labeling in 6-weeks-old male Thy1-GFP mice. As Thy1-GFP expression was restricted to maturated granule cells, it was combined with doublecortin-immunolabeling of immature granule cells. We developed a novel classification system that is easily applicable and enables objective and direct categorization of newborn granule cells based on the degree of dendritic development in relation to the layer specificity of the dentate gyrus. The structural development of adult-generated granule cells was correlated with age, albeit with notable differences in the time course of development between individual cells. In addition, the size of the nucleus, immunolabeled with the granule cell specific marker Prospero-related homeobox 1 gene, was a stable indicator of the degree of a cell's structural maturation and could be used as a straightforward parameter of granule cell development. Therefore, further studies could employ our doublecortin-staging system and nuclear size measurement to perform investigations of morphological development in combination with functional studies of adult-born granule cells. Furthermore, the Thy1-GFP transgenic mouse model can be used as an additional investigation tool because the reporter gene labels granule cells that are 4 weeks or older, while very young cells could be visualized through the immature marker doublecortin. This will enable comparison studies regarding the structure and function between young immature and older matured granule cells.
Background: From 2008–2013, the European indication for panitumumab required that patients' tumor KRAS exon 2 mutation status was known prior to starting treatment. To evaluate physician awareness of panitumumab prescribing information and how physicians prescribe panitumumab in patients with metastatic colorectal cancer (mCRC), two European multi-country, cross-sectional, observational studies were initiated in 2012: a physician survey and a medical records review. The first two out of three planned rounds for each study are reported.
Methods: The primary objective in the physician survey was to estimate the prevalence of KRAS testing, and in the medical records review, it was to evaluate the effect of test results on patterns of panitumumab use. The medical records review study also included a pathologists' survey.
Results: In the physician survey, nearly all oncologists (299/301) were aware of the correct panitumumab indication and the need to test patients' tumor KRAS status before treatment with panitumumab. Nearly all oncologists (283/301) had in the past 6 months of clinical practice administered panitumumab correctly to mCRC patients with wild-type KRAS status. In the medical records review, 97.5% of participating oncologists (77/79) conducted a KRAS test for all of their patients prior to prescribing panitumumab. Four patients (1.3%) did not have tumor KRAS mutation status tested prior to starting panitumumab treatment. Approximately one-quarter of patients (85/306) were treated with panitumumab and concurrent oxaliplatin-containing chemotherapy; of these, 83/85 had confirmed wild-type KRAS status prior to starting panitumumab treatment. All 56 referred laboratories that participated used a Conformité Européenne-marked or otherwise validated KRAS detection method, and nearly all (55/56) participated in a quality assurance scheme.
Conclusions: There was a high level of knowledge amongst oncologists around panitumumab prescribing information and the need to test and confirm patients' tumors as being wild-type KRAS prior to treatment with panitumumab, with or without concurrent oxaliplatin-containing therapy.
Magnetic resonance imaging (MRI) provides non-invasive, repetitive measures in the same individual, allowing the study of a physio-pathological event over time. In this study, we tested the performance of 7 Tesla multi-parametric MRI to monitor the dynamic changes of mouse skeletal muscle injury and regeneration upon acute ischemia induced by femoral artery dissection. T2-mapping (T2 relaxation time), diffusion-tensor imaging (Fractional Anisotropy) and perfusion by Dynamic Contrast-Enhanced MRI (K-trans) were measured and imaging results were correlated with histological morphometric analysis in both Gastrocnemius and Tibialis anterior muscles. We found that tissue damage positively correlated with T2-relaxation time, while myofiber regeneration and capillary density positively correlated with Fractional Anisotropy. Interestingly, K-trans positively correlated with capillary density. Accordingly, repeated MRI measurements between day 1 and day 28 after surgery in ischemic muscles showed that: 1) T2-relaxation time rapidly increased upon ischemia and then gradually declined, returning almost to basal level in the last phases of the regeneration process; 2) Fractional Anisotropy dropped upon ischemic damage induction and then recovered along with muscle regeneration and neoangiogenesis; 3) K-trans reached a minimum upon ischemia, then progressively recovered. Overall, Gastrocnemius and Tibialis anterior muscles displayed similar patterns of MRI parameters dynamic, with more marked responses and less variability in Tibialis anterior. We conclude that MRI provides quantitative information about both tissue damage after ischemia and the subsequent vascular and muscle regeneration, accounting for the differences between subjects and, within the same individual, between different muscles.
Epigenetic marks critically control gene expression and thus the cellular activity state. The functions of many epigenetic modifiers in the vascular system have not yet been studied. We screened for histone modifiers in endothelial cells and observed a fairly high expression of the histone plant homeodomain finger protein 8 (PHF8). Given its high expression, we hypothesize that this histone demethylase is important for endothelial cell function. Overexpression of PHF8 catalyzed the removal of methyl-groups from histone 3 lysine 9 (H3K9) and H4K20, whereas knockdown of the enzyme increased H3K9 methylation. Knockdown of PHF8 by RNAi also attenuated endothelial proliferation and survival. As a functional readout endothelial migration and tube formation was studied. PHF8 siRNA attenuated the capacity for migration and developing of capillary-like structures. Given the impact of PHF8 on cell cycle genes, endothelial E2F transcription factors were screened, which led to the identification of the gene repressor E2F4 to be controlled by PHF8. Importantly, PHF8 maintains E2F4 but not E2F1 expression in endothelial cells. Consistently, chromatin immunoprecipitation revealed that PHF8 reduces the H3K9me2 level at the E2F4 transcriptional start site, demonstrating a direct function of PHF8 in endothelial E2F4 gene regulation. Conclusion: PHF8 by controlling E2F4 expression maintains endothelial function.
Background: Chronic ethanol (EtOH) abuse worsens pathophysiological derangements after hemorrhagic shock and resuscitation (H/R) that induce hepatic injury and strong inflammatory changes via JNK and NF-κB activation. Inhibiting JNK with a cell-penetrating, protease-resistant peptide D-JNKI-1 after H/R in mice with healthy livers ameliorated these effects. Here, we studied if JNK inhibition by D-JNKI-1 in chronically EtOH-fed mice after hemorrhagic shock prior to the onset of resuscitation also confers protection.
Methods: Male mice were fed a Lieber-DeCarli diet containing EtOH or an isocaloric control (ctrl) diet for 4 weeks. Animals were hemorrhaged for 90 min (32 ± 2 mm Hg) and randomly received either D-JNKI-1 (11 mg/kg, intraperitoneally, i. p.) or sterile saline as vehicle (veh) immediately before the onset of resuscitation. Sham animals underwent surgical procedures without H/R and were either D-JNKI-1 or veh treated. Two hours after resuscitation, blood samples and liver tissue were harvested.
Results: H/R induced hepatic injury with increased systemic interleukin (IL)-6 levels, and enhanced local gene expression of NF-κB-controlled genes such as intercellular adhesion molecule (ICAM)-1 and matrix metallopeptidase (MMP)9. c-Jun and NF-κB phosphorylation were increased after H/R. These effects were further increased in EtOH-fed mice after H/R. D-JNKI-1 application inhibited the proinflammatory changes and reduced significantly hepatic injury after H/R in ctrl-fed mice. Moreover, D-JNKI-1 reduces in ctrl-fed mice the H/R-induced c-Jun and NF-κB phosphorylation. However, in chronically EtOH-fed mice, JNK inhibition did not prevent the H/R-induced hepatic damage and proinflammatory changes nor c-Jun and NF-κB phosphorylation after H/R.
Conclusions: These results indicate, that JNK inhibition is protective only in not pre-harmed liver after H/R. In contrast, the pronounced H/R-induced liver damage in mice being chronically fed with ethanol cannot be prevented by JNK inhibition after H/R and seems to be under the control of NF-κB.
Long-term potentiation (LTP) and long-term depression (LTD) are widely accepted to be synaptic mechanisms involved in learning and memory. It remains uncertain, however, which particular activity rules are utilized by hippocampal neurons to induce LTP and LTD in behaving animals. Recent experiments in the dentate gyrus of freely moving rats revealed an unexpected pattern of LTP and LTD from high-frequency perforant path stimulation. While 400 Hz theta-burst stimulation (400-TBS) and 400 Hz delta-burst stimulation (400-DBS) elicited substantial LTP of the tetanized medial path input and, concurrently, LTD of the non-tetanized lateral path input, 100 Hz theta-burst stimulation (100-TBS, a normally efficient LTP protocol for in vitro preparations) produced only weak LTP and concurrent LTD. Here we show in a biophysically realistic compartmental granule cell model that this pattern of results can be accounted for by a voltage-based spike-timing-dependent plasticity (STDP) rule combined with a relatively fast Bienenstock-Cooper-Munro (BCM)-like homeostatic metaplasticity rule, all on a background of ongoing spontaneous activity in the input fibers. Our results suggest that, at least for dentate granule cells, the interplay of STDP-BCM plasticity rules and ongoing pre- and postsynaptic background activity determines not only the degree of input-specific LTP elicited by various plasticity-inducing protocols, but also the degree of associated LTD in neighboring non-tetanized inputs, as generated by the ongoing constitutive activity at these synapses.
Leaky integrate-and-fire (LIF) network models are commonly used to study how the spiking dynamics of neural networks changes with stimuli, tasks or dynamic network states. However, neurophysiological studies in vivo often rather measure the mass activity of neuronal microcircuits with the local field potential (LFP). Given that LFPs are generated by spatially separated currents across the neuronal membrane, they cannot be computed directly from quantities defined in models of point-like LIF neurons. Here, we explore the best approximation for predicting the LFP based on standard output from point-neuron LIF networks. To search for this best “LFP proxy”, we compared LFP predictions from candidate proxies based on LIF network output (e.g, firing rates, membrane potentials, synaptic currents) with “ground-truth” LFP obtained when the LIF network synaptic input currents were injected into an analogous three-dimensional (3D) network model of multi-compartmental neurons with realistic morphology, spatial distributions of somata and synapses. We found that a specific fixed linear combination of the LIF synaptic currents provided an accurate LFP proxy, accounting for most of the variance of the LFP time course observed in the 3D network for all recording locations. This proxy performed well over a broad set of conditions, including substantial variations of the neuronal morphologies. Our results provide a simple formula for estimating the time course of the LFP from LIF network simulations in cases where a single pyramidal population dominates the LFP generation, and thereby facilitate quantitative comparison between computational models and experimental LFP recordings in vivo.
The flow-responsive transcription factor Krüppel-like factor 2 (KLF2) maintains an anti-coagulant, anti-inflammatory endothelium with sufficient nitric oxide (NO)-bioavailability. In this study, we aimed to explore, both in vitro and in human vascular tissue, expression of the NO-transporting transmembrane pore aquaporin-1 (AQP1) and its regulation by atheroprotective KLF2 and atherogenic inflammatory stimuli. In silico analysis of gene expression profiles from studies that assessed the effects of KLF2 overexpression in vitro and atherosclerosis in vivo on endothelial cells, identifies AQP1 as KLF2 downstream gene with elevated expression in the plaque-free vessel wall. Biomechanical and pharmaceutical induction of KLF2 in vitro is accompanied by induction of AQP1. Chromosome immunoprecipitation (CHIP) confirms binding of KLF2 to the AQP1 promoter. Inflammatory stimulation of endothelial cells leads to repression of AQP1 transcription, which is restrained by KLF2 overexpression. Immunohistochemistry reveals expression of aquaporin-1 in non-activated endothelium overlying macrophage-poor intimae, irrespective whether these intimae are characterized as being plaque-free or as containing advanced plaque. We conclude that AQP1 expression is subject to KLF2-mediated positive regulation by atheroprotective shear stress and is downregulated under inflammatory conditions both in vitro and in vivo. Thus, endothelial expression of AQP1 characterizes the atheroprotected, non-inflamed vessel wall. Our data provide support for a continuous role of KLF2 in stabilizing the vessel wall via co-temporal expression of eNOS and AQP1 both preceding and during the pathogenesis of atherosclerosis.
Different highly effective interferon-free treatment options for chronic hepatitis C virus (HCV) infection are currently available. Pre-existence of resistance associated variants (RAVs) to direct antiviral agents (DAAs) reduces sustained virologic response (SVR) rates by 3–53% in hepatitis C virus (HCV) genotype 1 infected patients depending on different predictors and the DAA regimen used. Frequencies of single and combined resistance to NS3, NS5A and NS5B inhibitors and consequences for the applicability of different treatment regimens are unknown. Parallel population based sequencing of HCV NS3, NS5A and NS5B genes in 312 treatment-naïve Caucasian HCV genotype 1 infected patients showed the presence of major resistant variants in 20.5% (NS3), 11.9% (NS5A), and 22.1% (NS5B) with important differences for HCV subtypes. In NS3, Q80K was observed in 34.7% and 2.1% of subtype 1a and 1b patients, respectively while other RAVs to second generation protease inhibitors were detected rarely (1.4%). Within NS5A RAVs were observed in 7.1% of subtype 1a and 17.6% in subtype 1b infected patients. RAVs to non-nucleoside NS5B inhibitors were observed in 3.5% and 44.4% of subtype 1a and 1b patients, respectively. Considering all three DAA targets all subtype 1a and 98.6% of subtype 1b infected patients were wildtype for at least one interferon free DAA regimen currently available. In conclusion, baseline resistance testing allows the selection of at least one RAVs-free treatment option for nearly all patients enabling a potentially cost- and efficacy-optimized treatment of chronic hepatitis C.
Introduction: Aim of this study was to reduce blood loss caused by diagnostic blood sampling and to minimize the development of anemia in a high-risk group of mechanically ventilated medical intensive care patients. We therefore implemented a “blood-saving bundle” (BSB) combining a closed-loop arterial blood sampling system, smaller sampling tubes, reduced frequency of blood drawings, and reduced sample numbers.
Methods: The study included all patients from our medical ICU who were ventilated for more than 72 hours. Exclusion criteria were: acute or chronic anemia on admission, bleeding episode(s) during the ICU stay, or end-of-life therapy. The BSB was introduced in 2009 with training and educational support. Patients treated in 2008, before the introduction of the BSB, served as a control group (n = 41, 617 observation days), and were compared with patients treated in 2010 after the introduction of the BSB (BSB group, n = 50, 559 observation days). Primary endpoints were blood loss per day, and development of anemia. Secondary endpoints were numbers of blood transfusions, number of days on mechanical ventilation, and length of the ICU stay.
Results: Mean blood loss per ICU day was decreased from 43.3 ml (95% CI: 41.2 to 45.3 ml) in the controls to 15.0 ml (14.3 to 15.7 ml) in the BSB group (P < 0.001). The introduction of a closed-loop arterial blood sampling system was the major contributor to this effect. Mean hemoglobin concentrations showed no significant differences in both groups during the ICU stay. Hemoglobin values <9 g/dl, however, were recorded in 21.2% of observation days in the controls versus 15.4% in the BSB group (P = 0.01). Units of transfused red blood cells per 100 observation days decreased from 7 to 2.3 (P < 0.001). The mean number of ventilation days was 7.1 days (6.1 to 8.3 days) in the controls and 7.5 days (6.6 to 8.5 days) in the BSB group (P = NS). In total, patients in the BSB group stayed in ICU for a mean of 9.9 days (8.6 to 11.3 days), compared to a mean ICU stay of 13.0 days (10.9 to 15.4 days) in the control group (P = 0.014). Due to the longitudinal study design, however, we cannot exclude uncontrolled confounders affecting the transfusion frequency and mean ICU stay.
Conclusion: Our BSB could be easily implemented and was able to reduce diagnostic blood loss.
Introduction: The Helicopter Emergency Medical Service (HEMS) was established for the prehospital trauma care of patients. Improved rescue times and increased coverage areas are discussed as specific advantages of HEMS. We recently found evidence that HEMS exerts beneficial effects on outcomes for severely injured patients. However, it still remains unknown which group of trauma patients might benefit most from HEMS rescue. Consequently, the unique aim of this study was to reveal which patients might benefit most from HEMS rescue.
Methods: Trauma patients (ISS ≥9) primarily treated by HEMS or ground emergency medical services (GEMS) between 2002 and 2012 were analysed using the TraumaRegister DGU. A multivariate regression analysis was used to reveal the survival benefit between different trauma populations.
Results: The study included 52 281 trauma patients. Of these, 68.8% (35 974) were rescued by GEMS and 31.2% (16 307) by HEMS. HEMS patients were more severely injured compared to GEMS patients (ISS: HEMS 24.8±13.5 vs. GEMS 21.7±18.0) and more frequently suffered traumatic shock (SBP sys <90mmHg: HEMS 18.3% vs. GEMS 14.8%). However, logistic regression analysis revealed that HEMS rescues resulted in an overall survival benefit compared to GEMS (OR 0.81, 95% CI [0.75–0.87], p<0.001, Nagelkerke's R squared 0.526, area under the ROC curve 0.922, 95% CI [0.919–0.925]). Analysis of specific subgroups demonstrated that patients aged older than 55 years (OR 0.62, 95% CI [0.50–0.77]) had the highest survival benefit after HEMS treatment. Furthermore, HEMS rescue had the most significant impact after ‘low falls’ (OR 0.68, 95% CI [0.55–0.84]) and in the case of minor severity injuries (ISS 9–15) (OR 0.66, 95% CI [0.49–0.88]).
Conclusions: In general, trauma patients benefit from HEMS rescue with in-hospital survival as the main outcome parameter. Focusing on special subgroups, middle aged and older patients, low-energy trauma, and minor severity injuries had the highest survival benefit when rescued by HEMS. Further studies are required to determine the potential reasons of this benefit.
Multiple antenatal dexamethasone treatment alters brain vessel differentiation in newborn mouse pups
(2015)
Antenatal steroid treatment decreases morbidity and mortality in premature infants through the maturation of lung tissue, which enables sufficient breathing performance. However, clinical and animal studies have shown that repeated doses of glucocorticoids such as dexamethasone and betamethasone lead to long-term adverse effects on brain development. Therefore, we established a mouse model for antenatal dexamethasone treatment to investigate the effects of dexamethasone on brain vessel differentiation towards the blood-brain barrier (BBB) phenotype, focusing on molecular marker analysis. The major findings were that in total brains on postnatal day (PN) 4 triple antenatal dexamethasone treatment significantly downregulated the tight junction protein claudin-5, the endothelial marker Pecam-1/CD31, the glucocorticoid receptor, the NR1 subunit of the N-methyl-D-aspartate receptor, and Abc transporters (Abcb1a, Abcg2 Abcc4). Less pronounced effects were found after single antenatal dexamethasone treatment and in PN10 samples. Comparisons of total brain samples with isolated brain endothelial cells together with the stainings for Pecam-1/CD31 and claudin-5 led to the assumption that the morphology of brain vessels is affected by antenatal dexamethasone treatment at PN4. On the mRNA level markers for angiogenesis, the sonic hedgehog and the Wnt pathway were downregulated in PN4 samples, suggesting fundamental changes in brain vascularization and/or differentiation. In conclusion, we provided a first comprehensive molecular basis for the adverse effects of multiple antenatal dexamethasone treatment on brain vessel differentiation.
Background: SNPs near the interferon lambda (IFNL) 3 gene are predictors for sustained virological response (SVR) in patients with chronic hepatitis C genotype (GT) 1. In addition, a dinucleotide frame shift in ss469415590 was described, which generates IFNL4. In this study, we compared the role of IFNL4 variants with IFNL3-(rs12979860) and IFNL3-(rs8099917) on response to pegylated (PEG)-IFN and Ribavirin (RBV) in patients with chronic hepatitis C GT2/3.
Methods: We recruited 1006 patients with chronic hepatitis C and GT2/3 in a large German registry. A treatment with PEG-IFN and Ribavirin was started by 959 patients. We performed genotyping of IFNL3 (rs12979860, n = 726; rs8099917, n = 687) and of IFNL4 (ss469415590; n = 631).
Results: Both preferable IFNL3 genotypes were associated with RVR (both p<0.0001) rather than with SVR (rs12979860: p = 0.251; rs8099917: p = 0.447). Only RVR was linked to SVR in univariate and multivariate analyzes (both p<0.001). Concordance of genotyping in patients with available serum samples and EDTA blood samples (n = 259) was more than 96% for both IFNL3 SNPs. IFNL3-(rs12979860) correlated with IFNL4: 99.2% of patients with IFNL3-(rs12979860)-CC were IFNL4-(ss469415590)-TT/TT. IFNL3-(rs12979860)-CT was linked with IFNL4-(ss469415590)-TT/ΔG (98.0%) and IFNL3-(rs12979860)-TT was associated with IFNL4-(ss469415590)-ΔG/ΔG (97.6%).
Conclusion: IFNL3 genotyping from serum was highly efficient and can be used as an alternative if EDTA whole blood is not available. In Caucasian GT2/3 patients genotyping for INFL4-(ss469415590) does not lead to additional information for the decision-making process. Importantly, IFNL3 SNPs were not associated with SVR but with RVR. Even in the era of new direct acting antiviral (DAA) therapies, IFNL3 testing may therefore still be considered for naïve GT2/3 patients to decide if dual Peg-IFN/RBV therapy is an option in resource limited regions.
Background & aim: Whether inosine triphosphatase (ITPA) gene polymorphisms predict anemia during interferon-free therapy in chronic hepatitis C virus (HCV)-infected patients is unknown. We examined the relationship between two ITPA polymorphisms, anemia, and sustained virological response 12 weeks post-treatment (SVR12) in patients receiving the NS3/4A protease inhibitor faldaprevir, the non-nucleoside polymerase inhibitor deleobuvir, and ribavirin.
Methods: HCV genotype 1-infected, treatment-naïve patients (N = 362) were randomized and treated in one of five treatment arms with faldaprevir and deleobuvir with or without ribavirin. Two ITPA polymorphisms (rs1127354 and rs6051702) were genotyped and defined as ITPA-deficient (rs1127354 AA or AC; rs6051702 CC or CA) or ITPA-non-deficient (rs1127354 CC; rs6051702 AA) according to their association with ITPA deficiency. Baseline and on-treatment variables associated with anemia and SVR12 were identified using logistic regression.
Results: In the pooled ribavirin-containing arms, 10.1% (32/316) of patients experienced on-treatment hemoglobin <10 g/dL, and 32.6% (103/316) experienced on-treatment hemoglobin <10 g/dL or a change from baseline ≥3.5 g/dL. Of the latter group, 99% (102/103) had the ITPA-non-deficient rs1127354 genotype. Other variables associated with on-treatment hemoglobin <10 g/dL or a decrease ≥3.5 g/dL were age, baseline hemoglobin, rs6051702 genotype, and plasma ribavirin concentration. In a multivariate analysis, high plasma ribavirin concentration, low baseline hemoglobin, HCV genotype 1b, and IL28B genotype CC were associated with higher SVR12.
Conclusions: The ITPA rs1127354 CC and rs6051702 AA genotypes may predict ribavirin-induced anemia during treatment with interferon-free, ribavirin-containing regimens. With this interferon-free regimen, SVR was associated with ribavirin levels, but not with anemia or ITPA genotypes.
Abstract: The current Ebola outbreak poses a threat to individual and global public health. Although the disease has been of interest to the scientific community since 1976, an effective vaccination approach is still lacking. This fact questions past global public health strategies, which have not foreseen the possible impact of this infectious disease. To quantify the global research activity in this field, a scientometric investigation was conducted. We analyzed the research output of countries, individual institutions and their collaborative networks. The resulting research architecture indicated that American and European countries played a leading role regarding output activity, citations and multi- and bilateral cooperations. When related to population numbers, African countries, which usually do not dominate the global research in other medical fields, were among the most prolific nations. We conclude that the field of Ebola research is constantly progressing, and the research landscape is influenced by economical and infrastructural factors as well as historical relations between countries and outbreak events.
Author Summary: For the first time in the history of the disease, the Ebola virus left its local setting and affected people not only in isolated rural areas, but reached larger towns and cities leading to worldwide repercussions. This development prompted a joint global response to this health threat. This encompassed not only immediate relief efforts, but also an up search in global research work. In this study, the scientific output in Ebola research available in one of the mayor medical search platforms was characterized. We studied among others the origin of research, the collaboration between countries and the research topics. Partly, the obtained data was weighted against economic parameters. We attained a detailed map of the research activities from the discovery of Ebola in 1976 up to today. Our research provides the first overview of the worldwide Ebola research output. It might help stakeholders in Ebola research to better plan investigations with a global perspective.
Pathogenicity of many microbes relies on their capacity to resist innate immunity, and to survive and persist in an immunocompetent human host microbes have developed highly efficient and sophisticated complement evasion strategies. Here we show that different human pathogens including Gram-negative and Gram-positive bacteria, as well as the fungal pathogen Candida albicans, acquire the human terminal complement regulator vitronectin to their surface. By using truncated vitronectin fragments we found that all analyzed microbial pathogens (n = 13) bound human vitronectin via the same C-terminal heparin-binding domain (amino acids 352–374). This specific interaction leaves the terminal complement complex (TCC) regulatory region of vitronectin accessible, allowing inhibition of C5b-7 membrane insertion and C9 polymerization. Vitronectin complexed with the various microbes and corresponding proteins was thus functionally active and inhibited complement-mediated C5b-9 deposition. Taken together, diverse microbial pathogens expressing different structurally unrelated vitronectin-binding molecules interact with host vitronectin via the same conserved region to allow versatile control of the host innate immune response.
Salt bridges in lipid bilayers play a decisive role in the dynamic assembly and downstream signaling of the natural killer and T-cell receptors. Here, we describe the identification of an inter-subunit salt bridge in the membrane within yet another key component of the immune system, the peptide-loading complex (PLC). The PLC regulates cell surface presentation of self-antigens and antigenic peptides via molecules of the major histocompatibility complex class I. We demonstrate that a single salt bridge in the membrane between the transporter associated with antigen processing TAP and the MHC I-specific chaperone tapasin is essential for the assembly of the PLC and for efficient MHC I antigen presentation. Molecular modeling and all-atom molecular dynamics simulations suggest an ionic lock-switch mechanism for the binding of TAP to tapasin, in which an unfavorable uncompensated charge in the ER-membrane is prevented through complex formation. Our findings not only deepen the understanding of the interaction network within the PLC, but also provide evidence for a general interaction principle of dynamic multiprotein membrane complexes in immunity.
Background: The treatment of different skin conditions with spa waters is a long tradition dating back to at least late Hellenism. Interestingly, independent scientific examinations studying the effect of spa waters are scarce.
Objective: In the present in vitro study, we compared the effect of culture media supplemented with (a) thermal spa waters (La Roche-Posay, Avène) and (b) two natural mineral drinking waters (Heppinger, Adelholzener) on physiological parameters in HaCaT keratinocytes.
Methods: The different medium preparations were investigated with regard to cell proliferation and cell damage. Moreover, the impact on inflammation parameters with and without ultraviolet B (UVB) irradiation was examined.
Results: Two popular thermal spring waters were found to suppress cell proliferation and cell damage. Moreover, these waters reversed the induction of interleukin-6, as measured using enzyme-linked immunosorbent assay and promoter transactivation, and the formation of reactive oxygen species after UVB stimulation. Of note, the two natural mineral waters, which are distributed as drinking waters, had some effect on the above-mentioned parameters but to a lesser extent.
Conclusion: In summary, our results show that spa waters, and particularly those derived from thermal springs, reduce parameters associated with inflammation. It seems likely that trace elements such as selenium and zinc are critical for the observed effects.
Emotional instability, difficulties in social adjustment, and disinhibited behavior are the most common symptoms of the psychiatric comorbidities in juvenile myoclonic epilepsy (JME). This psychopathology has been associated with dysfunctions of mesial-frontal brain circuits. The present work is a first direct test of this link and adapted a paradigm for probing frontal circuits during empathy for pain. Neural and psychophysiological parameters of pain empathy were assessed by combining functional magnetic resonance imaging (fMRI) with simultaneous pupillometry in 15 JME patients and 15 matched healthy controls. In JME patients, we observed reduced neural activation of the anterior cingulate cortex (ACC), the anterior insula (AI), and the ventrolateral prefrontal cortex (VLPFC). This modulation was paralleled by reduced pupil dilation during empathy for pain in patients. At the same time, pupil dilation was positively related to neural activity of the ACC, AI, and VLPFC. In JME patients, the ACC additionally showed reduced functional connectivity with the primary and secondary somatosensory cortex, areas fundamentally implicated in processing the somatic cause of another's pain. Our results provide first evidence that alterations of mesial-frontal circuits directly affect psychosocial functioning in JME patients and draw a link of pupil dynamics with brain activity during emotional processing. The findings of reduced pain empathy related activation of the ACC and AI and aberrant functional integration of the ACC with somatosensory cortex areas provide further evidence for this network's role in social behavior and helps explaining the JME psychopathology and patients' difficulties in social adjustment.
Introduction: For resectable soft tissue sarcoma (STS), radical surgery, usually combined with radiotherapy, is the mainstay of treatment and the only potentially curative modality. Since surgery is often complicated by large tumour size and extensive tumour vasculature, preoperative treatment strategies with the aim of devitalising the tumour are being explored. One option is treatment with antiangiogenic drugs. The multikinase inhibitor pazopanib, which possesses pronounced antiangiogenic effects, has shown activity in metastatic and unresectable STS, but has so far not been tested in the preoperative setting.
Methods and analysis: This open-label, multicentre phase II window-of-opportunity trial assesses pazopanib as preoperative treatment of resectable STS. Participants receive a 21-day course of pazopanib 800 mg daily during wait time for surgery. Major eligibility criteria are resectable, high-risk adult STS of any location, or metachronous solitary STS metastasis for which resection is planned, and adequate organ function and performance status. The trial uses an exact single-stage design. The primary end point is metabolic response rate (MRR), that is, the proportion of patients with >50% reduction of the mean standardised uptake value (SUVmean) in post-treatment compared to pre-treatment fluorodeoxyglucose positron emission tomography CT. The MRR below which the treatment is considered ineffective is 0.2. The MRR above which the treatment warrants further exploration is 0.4. With a type I error of 5% and a power of 80%, the sample size is 35 evaluable patients, with 12 or more responders as threshold. Main secondary end points are histopathological and MRI response, resectability, toxicity, recurrence-free and overall survival. In a translational substudy, endothelial progenitor cells and vascular epithelial growth factor receptor are analysed as potential prognostic and predictive markers.
Ethics and dissemination: Approval by the ethics committee II, University of Heidelberg, Germany (2012-019F-MA), German Federal Institute for Drugs and Medical Devices (61-3910-4038155) and German Federal Institute for Radiation Protection (Z5-22463/2-2012-007).
Trial registration number: NCT01543802, EudraCT: 2011-003745-18; Pre-results.
Effect of M3 muscarinic acetylcholine receptor deficiency on collagen antibody-induced arthritis
(2016)
Background: There is increasing evidence that the non-neuronal cholinergic system might be of importance for the pathology of rheumatoid arthritis. The role of M3 muscarinic acetylcholine receptor (M3R) in this regard has, however, not been investigated to date. Thus, in the present study we analyzed if M3R deficiency might have a protective effect on experimentally induced arthritis.
Methods: Collagen antibody-induced arthritis (CAIA) was evoked in M3R-deficient (M3R −/− ) mice and wild-type (WT) littermates. Severity of arthritis was assessed by scoring of paw swelling. The joints of arthritic and nonarthritic animals were analyzed for histopathological changes regarding synovial tissue, cartilage degradation and bone destruction. Further, gene expression analysis of respective markers was performed. Systemic and local inflammatory response was determined by flow cytometry and immunohistochemistry for leukocytes as well as mRNA and protein measurements for pro-inflammatory cytokines and chemokines.
Results: In arthritic M3R −/− mice the number of leukocytes, specifically neutrophils, was enhanced even though clinical arthritis score was not significantly different between WT and M3R −/− mice with CAIA. In M3R −/− mice, levels of neutrophil chemoattractant chemokine C-X-C-motif ligand 2 (CXCL2) as well as the pro-inflammatory cytokine interleukin-6 were already strongly increased in mice with low arthritis score, whereas WT mice only showed prominent expression of these markers when reaching high arthritis scores. Furthermore, arthritic M3R −/− mice displayed a stronger degradation of collagen II in the articular cartilage and, most strikingly, histopathological evaluation revealed more severe bone destruction in arthritic mice with M3R deficiency compared to WT littermates. Moreover, in M3R −/− mice, gene expression of markers for bone degradation (matrix metalloproteinase 13, cathepsin K and receptor activator of nuclear factor-κB ligand) was already increased in mice with low arthritis score.
Conclusions: Taken together, the present study shows that while M3R −/− mice were not protected from CAIA, they had a tendency toward a higher inflammatory response after arthritis induction than WT mice. Further, arthritis-induced joint destruction was significantly stronger in mice with M3R deficiency, indicating that stimulation of M3R might have protective effects on arthritis.
Background: Sphingolipids constitute bioactive molecules with functional implications in liver homeostasis. Particularly, ablation of very long chain ceramides in a knockout mouse model has been shown to cause a severe hepatopathy.
Methods: We aimed to evaluate the serum sphingolipid profile of 244 patients with cirrhosis prospectively followed for a median period of 228±217 days via mass spectrometry.
Results: We thereby observed a significant decrease of long and very long chain ceramides, particularly of C24ceramide, in patients with increasing severity of cirrhosis (p<0.001). Additionally, hydropic decompensation, defined by clinical presentation of ascites formation, was significantly correlated to low C24ceramide levels (p<0.001) while a significant association to hepatic decompensation and poor overall survival was observed for low serum concentrations of C24ceramide (p<0.001) as well. Multivariate analysis further identified low serum C24ceramide to be independently associated to overall survival (standard beta = -0.001, p = 0.022).
Conclusions: In our current analysis serum levels of very long chain ceramides show a significant reciprocal correlation to disease severity and hepatic decompensation and are independently associated with overall survival in patients with cirrhosis. Serum sphingolipid metabolites and particularly C24ceramide may constitute novel molecular targets of disease severity, hepatic decompensation and overall prognosis in cirrhosis and should be further evaluated in basic research studies.
Background & aims: Current guidelines recommend immunosuppressive treatment (IT) in patients with primary sclerosing cholangitis (PSC) and elevated aminotransferase levels more than five times the upper limit of normal and elevated serum IgG-levels above twice the upper limit of normal. Since there is no evidence to support this recommendation, we aimed to assess the criteria that guided clinicians in clinical practice to initiate IT in patients with previously diagnosed PSC.
Methods: This is a retrospective analysis of 196 PSC patients from seven German hepatology centers, of whom 36 patients had received IT solely for their liver disease during the course of PSC. Analyses were carried out using methods for competing risks.
Results: A simplified autoimmune hepatitis (AIH) score >5 (HR of 36, p<0.0001) and a modified histological activity index (mHAI) greater than 3/18 points (HR 3.6, p = 0.0274) were associated with the initiation of IT during the course of PSC. Of note, PSC patients who subsequently received IT differed already at the time of PSC diagnosis from those patients, who did not receive IT during follow-up: they presented with increased levels of IgG (p = 0.004) and more frequently had clinical signs of cirrhosis (p = 0.0002).
Conclusions: This is the first study which investigates the parameters associated with IT in patients with PSC in clinical practice. A simplified AIH score >5 and a mHAI score >3, suggesting concomitant features of AIH, influenced the decision to introduce IT during the course of PSC. In German clinical practice, the cutoffs used to guide IT may be lower than recommended by current guidelines.
Introduction: The effects of manipulated dental occlusion on body posture has been investigated quite often and discussed controversially in the literature. Far less attention has been paid to the influence of dental occlusion position on human movement. If human movement was analysed, it was mostly while walking and not while running. This study was therefore designed to identify the effect of lower jaw positions on running behaviour according to different dental occlusion positions.
Methods: Twenty healthy young recreational runners (mean age = 33.9±5.8 years) participated in this study. Kinematic data were collected using an eight-camera Vicon motion capture system (VICON Motion Systems, Oxford, UK). Subjects were consecutively prepared with four different dental occlusion conditions in random order and performed five running trials per test condition on a level walkway with their preferred running shoes. Vector based pattern recognition methods, in particular cluster analysis and support vector machines (SVM) were used for movement pattern identification.
Results: Subjects exhibited unique movement patterns leading to 18 clusters for the 20 subjects. No overall classification of the splint condition could be observed. Within individual subjects different running patterns could be identified for the four splint conditions. The splint conditions lead to a more symmetrical running pattern than the control condition.
Discussion: The influence of an occlusal splint on running pattern can be confirmed in this study. Wearing a splint increases the symmetry of the running pattern. A more symmetrical running pattern might help to reduce the risk of injuries or help in performance. The change of the movement pattern between the neutral condition and any of the three splint conditions was significant within subjects but not across subjects. Therefore the dental splint has a measureable influence on the running pattern of subjects, however subjects individuality has to be considered when choosing the optimal splint condition for a specific subject.
Objectives: We aimed to investigate the contemporary usage rate and habits of the WHO Surgical Safety Checklist (SSC) in German urological departments.
Methods: We designed a 26-item questionnaire that was sent to all urological departments in Germany. The primary aim of this study was to evaluate the usage rate of the SSC. Secondary aims were to compare perioperative characteristics of users vs. non-users of the SSC and to assess circumstances of the SSC application.
Results: A total of 213 of 234 (91 %) urological departments were users of the SSC, and 21 (9 %) were non-users. SSC users had more often a standard protocol, took less time and had fewer people involved for checking perioperative patient data compared to non-users. Financial budgeting for the SSC existed in 55 (24 %) departments and for patient safety in 73 (32 %) departments.
Conclusions: The usage rate of the SSC in urological departments in Germany is high despite restricted financial budgeting. Users of the SSC profit by saving time and manpower for checking perioperative patient data.
The purpose of this phase III clinical trial was to compare two different extracellular contrast agents, 1.0 M gadobutrol and 0.5 M gadopentate dimeglumine, for magnetic resonance imaging (MRI) in patients with known or suspected focal renal lesions. Using a multicenter, single-blind, interindividual, randomized study design, both contrast agents were compared in a total of 471 patients regarding their diagnostic accuracy, sensitivity, and specificity to correctly classify focal lesions of the kidney. To test for noninferiority the diagnostic accuracy rates for both contrast agents were compared with CT results based on a blinded reading. The average diagnostic accuracy across the three blinded readers (‘average reader’) was 83.7% for gadobutrol and 87.3% for gadopentate dimeglumine. The increase in accuracy from precontrast to combined precontrast and postcontrast MRI was 8.0% for gadobutrol and 6.9% for gadopentate dimeglumine. Sensitivity of the average reader was 85.2% for gadobutrol and 88.7% for gadopentate dimeglumine. Specificity of the average reader was 82.1% for gadobutrol and 86.1% for gadopentate dimeglumine. In conclusion, this study documents evidence for the noninferiority of a single i.v. bolus injection of 1.0 M gadobutrol compared with 0.5 M gadopentate dimeglumine in the diagnostic assessment of renal lesions with CE-MRI.