Die Bedeutung von Cofilin1 und Aktindynamik für die Funktion von T-Zellen

In der vorliegenden Arbeit wurde die Bedeutung des Aktin depolymerisierenden Faktors Cofilin1 und der Aktindynamik für die Funktion von T-Zellen näher untersucht. Der vollständige Knock-out des Cofilin1-Proteins führt zu früher embryonaler Letalität, sodass diese Problematik für die Untersuchungen umgangen werden musste. Dies erfolgte über einen molekulargenetischen Ansatz durch den Einsatz verschiedener Mausmodelle. Zunächst wurde über einen Tripletaustausch im Cofilin1-Gen konstitutiv aktives bzw. inaktives Protein hergestellt, um die Auswirkungen des Aktivierungszustands von Cofilin1 auf die Aktinpolymerisation zu untersuchen. In Aktin-Assays konnte bestätigt werden, dass konstitutiv aktives Cofilin1 eine vergleichbare Aktivität gegenüber Aktin zeigt wie das WT-Protein, während die inaktive Form keinen Einfluss nimmt. Anschließend wurden diese Mutationen in vivo im Mausmodell bezüglich ihrer Bedeutung in T-Zellen untersucht. Sowohl T-Helferzellen als auch zytotoxische T-Zellen mit konstitutiv aktivem Cofilin1 reifen im Thymus heran und wandern in die sekundären lymphatischen Organe aus. Das inaktive Cofilin1 führt dagegen, ebenso wie der frühe Knock-out des Proteins, zu einer Blockade in der T-Zellentwicklung. Des Weiteren wurde mittels Cofilin1-Deletion während einer späten Phase der T-Zellentwicklung die Bedeutung des Proteins in reifen T-Zellen untersucht. Es konnte gezeigt werden, dass eine Deletion des Cofilin1-Proteins zwar keinen Einfluss auf die T-Zellreifung hat, aber die T-Zellen nur zu einem kleinen Teil aus dem Thymus emigrieren können. Analysen von T-Zellen aus den sekundären lymphatischen Organen, wie Lymphknoten und Milz, machten eine starke Reduktion von T-Helferzellen und zytotoxischen T-Zellen mit Cofilin1-Defizienz in diesen Organen deutlich. Diese Zellen weisen ein erhöhtes Vorkommen von Apoptose auf. Das Fehlen von Cofilin1-defizienten T-Zellen ist vermutlich auf Migrations- und Adhäsionsdefekte zurückzuführen. Detaillierte Untersuchungen bezüglich der Aktivierbarkeit von zytotoxischen T-Zellen mit Cofilin1-Knock-out konnten belegen, dass sie hinsichtlich dieses Aspekts einen Defekt aufweisen. Weiterhin stellte die Aptamer-Technologie einen zweiten Ansatz zur spezifischen Regulation der Cofilin1-Aktivität dar. Es konnte erfolgreich ein RNA-Aptamer selektiert werden, welches eine Affinität für Cofilin1 besitzt

CO-PARENTING FACTORS THAT CONTRIBUTE TO ACADEMIC SUCCESS

This study assessed factors contributed from parents who live in two different households and that lead to academic success. Data were collected from undergraduates enrolled in a Midwestern satellite university. Academic success was defined by university enrollment, grade point average, and standardized testing scores. Co-parenting factors that were hypothesized to lead to academic success included the distance between parents homes (which further influenced time spent with the child, participation in child‘s activities, and participation in decision making) and financial stability (which also influenced participation in decision making and the level of conflict within the family). The original structural equation model revealed that the relationship linking the distance between homes and the time spent with the child was accurately described. Added to the model, after the Lagrange test, was a path from finances to participation in child‘s activities and time spent with the child. The financial stability of a family predicted the participation of the non-custodial parent in the child‘s activities, in the decision-making for the child, conflict, and the time spent with the child. Implications for practitioners who work with families with co-parenting responsibilities are discussed

CO-PARENTING FACTORS THAT CONTRIBUTE TO ACADEMIC SUCCESS

This study assessed factors contributed from parents who live in two different households and that lead to academic success. Data were collected from undergraduates enrolled in a Midwestern satellite university. Academic success was defined by university enrollment, grade point average, and standardized testing scores. Co-parenting factors that were hypothesized to lead to academic success included the distance between parents homes (which further influenced time spent with the child, participation in child‘s activities, and participation in decision making) and financial stability (which also influenced participation in decision making and the level of conflict within the family). The original structural equation model revealed that the relationship linking the distance between homes and the time spent with the child was accurately described. Added to the model, after the Lagrange test, was a path from finances to participation in child‘s activities and time spent with the child. The financial stability of a family predicted the participation of the non-custodial parent in the child‘s activities, in the decision-making for the child, conflict, and the time spent with the child. Implications for practitioners who work with families with co-parenting responsibilities are discussed

The effect of CpG-ODN on antigen presenting cells of the foal

BACKGROUND: Cytosine-phosphate-guanosine oligodeoxynucleotide (CpG-ODN) has been used successfully to induce immune responses against viral and intracellular organisms in mammals. The main objective of this study was to test the effect of CpG-ODN on antigen presenting cells of young foals. METHODS: Peripheral blood monocytes of foals (n = 7) were isolated in the first day of life and monthly thereafter up to 3 months of life. Adult horse (n = 7) monocytes were isolated and tested once for comparison. Isolated monocytes were stimulated with IL-4 and GM-CSF (to obtain dendritic cells, DC) or not stimulated (to obtain macrophages). Macrophages and DCs were stimulated for 14-16 hours with either CpG-ODN, LPS or not stimulated. The stimulated and non-stimulated cells were tested for cell surface markers (CD86 and MHC class II) using flow cytometry, mRNA expression of cytokines (IL-12, IFNalpha, IL-10) and TLR-9 using real time quantitative RT-PCR, and for the activation of the transcription factor NF-kappaB p65 using a chemiluminescence assay. RESULTS: The median fluorescence of the MHC class II molecule in non-stimulated foal macrophages and DCs at birth were 12.5 times and 11.2 times inferior, respectively, than adult horse cells (p = 0.009). That difference subsided at 3 months of life (p = 0.3). The expression of the CD86 co-stimulatory molecule was comparable in adult horse and foal macrophages and DCs, independent of treatment. CpG-ODN stimulation induced IL-12p40 (53 times) and IFNalpha (23 times) mRNA expression in CpG-ODN-treated adult horse DCs (p = 0.078), but not macrophages, in comparison to non-stimulated cells. In contrast, foal APCs did not respond to CpG-ODN stimulation with increased cytokine mRNA expression up to 3 months of age. TLR-9 mRNA expression and NF-kB activation (NF-kB p65) in foal DCs and macrophages were comparable (p \u3e 0.05) to adult horse cells. CONCLUSION: CpG-ODN treatment did not induce specific maturation and cytokine expression in foal macrophages and DCs. Nevertheless, adult horse DCs, but not macrophages, increased their expression of IL-12 and IFNalpha cytokines upon CpG-ODN stimulation. Importantly, foals presented an age-dependent limitation in the expression of MHC class II in macrophages and DCs, independent of treatment

Clinical outcomes of subcutaneous vs. transvenous implantable defibrillator therapy in a polymorbid patient cohort

BackgroundThe subcutaneous implantable cardioverter-defibrillator (S-ICD) has been designed to overcome lead-related complications and device endocarditis. Lacking the ability for pacing or resynchronization therapy its usage is limited to selected patients at risk for sudden cardiac death (SCD). ObjectiveThe aim of this single-center study was to assess clinical outcomes of S-ICD and single-chamber transvenous (TV)-ICD in an all-comers population. MethodsThe study cohort comprised a total of 119 ICD patients who underwent either S-ICD (n = 35) or TV-ICD (n = 84) implantation at the University Hospital Frankfurt from 2009 to 2017. By applying an inverse probability-weighting (IPW) analysis based on the propensity score including the Charlson Comorbidity Index (CCI) to adjust for potential extracardiac comorbidities, we aimed for head-to-head comparison on the study composite endpoint: overall survival, hospitalization, and device-associated events (including appropriate and inappropriate shocks or system-related complications). ResultsThe median age of the study population was 66.0 years, 22.7% of the patients were female. The underlying heart disease was ischemic cardiomyopathy (61.4%) with a median LVEF of 30%. Only 52.9% had received an ICD for primary prevention, most of the patients (67.3%) had advanced heart failure (NYHA class II-III) and 16.8% were in atrial fibrillation. CCI was 5 points in TV-ICD patients vs. 4 points for patients with S-ICD (p = 0.209) indicating increased morbidity. The composite endpoint occurred in 38 patients (31.9 %), revealing no significant difference between patients implanted with an S-ICD or TV-ICD (unweighted HR 1.50, 95 % confidence interval (CI) 0.78-2.90; p = 0.229, weighted HR 0.94, 95% CI, 0.61-1.50, p = 0.777). Furthermore, we observed no difference in any single clinical endpoint or device-associated outcome, neither in the unweighted cohort nor following inverse probability-weighting. ConclusionClinical outcomes of the S-ICD and TV-ICD revealed no differences in the composite endpoint including survival, freedom of hospitalization and device-associated events, even after careful adjustment for potential confounders. Moreover, the CCI was evaluated in a S-ICD cohort demonstrating higher survival rates than predicted by the CCI in young, polymorbid (S-)ICD patients

Systemic Effects by Intrathecal Administration of Triamcinolone Acetonide in Patients With Multiple Sclerosis

In patients suffering from multiple sclerosis (MS), intrathecal injection of triamcinolone acetonide (TCA) has been shown to improve symptoms of spasticity. Although repeated intrathecal injection of TCA has been used in a number of studies in late-stage MS patients with spinal cord involvement, no clinical-chemical data are available on the distribution of TCA in cerebrospinal fluid (CSF) or serum. Moreover, the effects of intrathecal TCA administration on the concentrations of endogenous steroids remain poorly understood. Therefore, we have quantified TCA and selected endogenous steroids in CSF and serum of TCA-treated MS patients suffering from spasticity. Concentrations of steroids were quantified by LC-MS, ELISA, or ECLIA and compared with the blood-brain barrier status, diagnosed with the Reibergram. The concentration of TCA in CSF significantly increased during each treatment cycle up to >5 mu g/ml both in male and female patients (p30 ng/ml (p< 0.001) and severely depressed serum levels of cortisol and corticosterone (p< 0.001). In addition, concentrations of circulating estrogen were significantly suppressed (p< 0.001). Due to the potent suppressive effects of TCA on steroid hormone concentrations both in the brain and in the periphery, we recommend careful surveillance of adrenal function following repeated intrathecal TCA injections in MS patients

Guideline for Trustworthy Artificial Intelligence -- AI Assessment Catalog

Artificial Intelligence (AI) has made impressive progress in recent years and represents a key technology that has a crucial impact on the economy and society. However, it is clear that AI and business models based on it can only reach their full potential if AI applications are developed according to high quality standards and are effectively protected against new AI risks. For instance, AI bears the risk of unfair treatment of individuals when processing personal data e.g., to support credit lending or staff recruitment decisions. The emergence of these new risks is closely linked to the fact that the behavior of AI applications, particularly those based on Machine Learning (ML), is essentially learned from large volumes of data and is not predetermined by fixed programmed rules. Thus, the issue of the trustworthiness of AI applications is crucial and is the subject of numerous major publications by stakeholders in politics, business and society. In addition, there is mutual agreement that the requirements for trustworthy AI, which are often described in an abstract way, must now be made clear and tangible. One challenge to overcome here relates to the fact that the specific quality criteria for an AI application depend heavily on the application context and possible measures to fulfill them in turn depend heavily on the AI technology used. Lastly, practical assessment procedures are needed to evaluate whether specific AI applications have been developed according to adequate quality standards. This AI assessment catalog addresses exactly this point and is intended for two target groups: Firstly, it provides developers with a guideline for systematically making their AI applications trustworthy. Secondly, it guides assessors and auditors on how to examine AI applications for trustworthiness in a structured way

Clonal hematopoiesis as a pitfall in germline variant interpretation in the context of Mendelian disorders

Clonal hematopoiesis because of somatic mutations in hematopoietic stem/progenitor cells is an age-related phenomenon and commonly observed when sequencing blood DNA in elderly individuals. Several genes that are implicated in clonal hematopoiesis are also associated with Mendelian disorders when mutated in the germline, potentially leading to variant misinterpretation. We performed a literature search to identify genes associated with age-related clonal hematopoiesis followed by an OMIM query to identify the subset of genes in which germline variants are associated with Mendelian disorders. We retrospectively screened for diagnostic cases in which the presence of age-related clonal hematopoiesis confounded exome sequencing data interpretation. We found 58 genes in which somatic mutations are implicated in clonal hematopoiesis, while germline variants in the same genes are associated with Mendelian (mostly neurodevelopmental) disorders. Using five selected cases of individuals with suspected monogenic disorders, we illustrate how clonal hematopoiesis in either variant databases or exome sequencing datasets poses a pitfall, potentially leading to variant misclassification and erroneous conclusions regarding gene-disease associations

Adaptive immune response to lipoproteins of Staphylococcus aureus in healthy subjects

Staphylococcus aureus is a frequent commensal but also a dangerous pathogen, causing many forms of infection ranging from mild to life-threatening conditions. Among its virulence factors are lipoproteins, which are anchored in the bacterial cell membrane. Lipoproteins perform various functions in colonization, immune evasion, and immunomodulation. These proteins are potent activators of innate immune receptors termed Toll-like receptors 2 and 6. This study addressed the specific B-cell and T-cell responses directed to lipoproteins in human S. aureus carriers and non-carriers. 2D immune proteomics and ELISA approaches revealed that titers of antibodies (IgG) binding to S. aureus lipoproteins were very low. Proliferation assays and cytokine profiling data showed only subtle responses of T cells; some lipoproteins did not elicit proliferation. Hence, the robust activation of the innate immune system by S. aureus lipoproteins does not translate into a strong adaptive immune response. Reasons for this may include inaccessibility of lipoproteins for B cells as well as ineffective processing and presentation of the antigens to T cells.</p