Analysis of sAC expression and function as a co-factor of CREB

Die lösliche Adenylatcyclase (sAC) katalysiert die Generierung von cAMP in speziellen Mikrodomänen im Zytoplasma und im Zellkern. In dieser Arbeit wurde eine Beteiligung der sAC an der Expression von cAMP-regulierten Genen, wie des epithelialen Na+ Kanals (ENaC) und der Na+/K+-ATPase, die beide den Aldosteron induzierten Na+ Strom über die Zellmembran vermitteln, gezeigt. Mittels Co-IP, ChIP und real time PCR wurde die sAC als Co-Faktor von CREB identifiziert, der direkt an DNA bindet und einen regulativen Einfluss auf die Aldosteron-vermittelte Genexpression ausübt. Zwei unabhängige Promotorregionen, die die Transkription der sAC zelltyp- und differenzierungsspezifisch steuern, wurden mittels Reportergenassay identifiziert. 4 kb der sAC 5´-flankierenden Region wurden in 60 CVD Patienten sequenziert und 7 genetische Varianten gefunden, wovon drei in einem Kopplungsungleichgewicht vorlagen und zwei molekulare Haplotypen mit Allel-spezifischer transkriptioneller Aktivität bilden. The soluble adenylyl cyclase (sAC) catalyzes the generation of cAMP in specified microdomains of the cytoplasm and the nucleus. In the current study it was demonstrated that sAC is involved in the expression of cAMP-regulated genes such as the epithelial Na+ channel (ENaC) and the Na+/K+-ATPase, both mediating the aldosterone-induced increase of Na+ currents across the cell membrane. Using Co-IP, ChIP and real time PCR sAC was identified as a co-factor of CREB, binding directly to DNA with regulative impact in aldosterone-mediated gene expression. Two distinct promoter regions to drive sAC transcription in a cell type- and differentiation-specific manner were identified using reportergene assay. Screening of 4 kb of the sAC 5'-flanking region in 60 patients with cardiovascular disease revealed seven genetic variants, three of which are in a strong linkage disequilibrium resulting in two molecular haplotypes with allele-specific transcriptional activity

Comparing a mindfulness- and CBT-based guided self-help Internet- and mobile-based intervention against a waiting list control condition as treatment for adults with frequent cannabis use: a randomized controlled trial of CANreduce 3.0

Background: Though Internet- and mobile-based interventions (IMIs) and mindfulness-based interventions (generally delivered in-situ) appear effective for people with substance use disorders, IMIs incorporating mindfulness are largely missing, including those targeting frequent cannabis use. Methods: This paper details the protocol for a three-arm randomized controlled trial comparing a mindfulness-based self-help IMI (arm 1) and cognitive-behavioral therapy (CBT)-based self-help IMI (arm 2) versus being on a waiting list (arm 3) in their effectiveness reducing cannabis use in frequent cannabis users. Predictors of retention, adherence and treatment outcomes will be identified and similarities between the two active intervention arms explored. Both active interventions last six weeks and consist of eight modules designed to reduce cannabis use and common mental health symptoms. With a targeted sample size of n = 210 per treatment arm, data will be collected at baseline immediately before program use is initiated; at six weeks, immediately after program completion; and at three and six months post baseline assessment to assess the retention of any gains achieved during treatment. The primary outcome will be number of days of cannabis use over the preceding 30 days. Secondary outcomes will include further measures of cannabis use and use of other substances, changes in mental health symptoms and mindfulness, client satisfaction, intervention retention and adherence, and adverse effects. Data analysis will follow ITT principles and primarily employ (generalized) linear mixed models. Discussion: This RCT will provide important insights into the effectiveness of an IMI integrating mindfulness to reduce cannabis use in frequent cannabis users. Trial registration: International Standard Randomized Controlled Trial Number Registry: ISRCTN14971662 ; date of registration: 09/09/2021. Keywords: Cannabis; Cognitive-behavioral therapy; Internet-based intervention; Mindfulness; Randomized controlled trial; Self-help

Detecting Bacteria on Wounds with Hyperspectral Imaging in Fluorescence Mode

Chronic non-healing wounds represent an increasing problem. In order to enable physicians and nurses to make evidence based decisions on wound treatment, the professional societies call for supporting tools to be offered to physicians. Oxygen supply, bacteria colonization and other parameters influence the healing process. So far, these parameters cannot be monitored in an objective and routinely manner. Existing methods like the microbiological analysis of wound swabs, mean a great deal of effort and partly a long delay. In this paper 42 fluorescence images from 42 patients with diabetic foot ulcer, recorded with a hyperspectral imaging system (TIVITA®), converted for fluorescence imaging, were analysed. Beside the fluorescence images, information about the bacterial colonization is available from microbiological analysis of wound swabs. After preprocessing, principal component analysis, PCA, is used for data analysis with a 405 nm excitation wavelength, the emission wavelength range 510 - 745 nm is used for analysis. After dividing the data into a training and a test dataset it could be shown, that bacteria are detectable in the wound area. A quantification in bacterial colonization counts (BCC) was not in the focus of the research in this study stage

Monitoring the electroactive cargo of extracellular vesicles can differentiate various cancer cell lines

Extracellular vesicles (EVs) are pivotal in cell-to-cell communication due to the array of cargo contained within these vesicles. EVs are considered important biomarkers for identification of disease, however most measurement approaches have focused on monitoring specific surface macromolecular targets. Our study focuses on exploring the electroactive component present within cargo from EVs obtained from various cancer and non-cancer cell lines using a disk carbon fiber microelectrode. Variations in the presence of oxidizable components were observed when the total cargo from EVs were measured, with the highest current detected in EVs from MCF7 cells. There were differences observed in the types of oxidizable species present within EVs from MCF7 and A549 cells. Single entity measurements showed clear spikes due to the detection of oxidizable cargo within EVs from MCF7 and A549 cells. These studies highlight the promise of monitoring EVs through the presence of varying electroactive components within the cargo and can drive a wave of new strategies towards specific detection of EVs for diagnosis and prognosis of various diseases

Was tun? Perspektiven für eine Unterrichtsqualitätsforschung der Zukunft

Die Unterrichtsqualitätsforschung hat in den vergangenen Jahrzehnten viel zum Verständnis der Merkmale beitragen, die einen qualitätsvollen Unterricht auszeichnen. Sie hat einflussreiche Modelle hervorgebracht, die eine gemeinsame Grundlage für die Erforschung des Unterrichts bereitstellen. Gleichzeitig bringen gesellschaftliche Veränderungsprozesse eine Neuorientierung des schulischen Unterrichts mit sich, und es lässt sich fragen, ob die Unterrichtsqualitätsforschung mit ihren bisherigen Ansätzen in der Lage ist, in Zukunft zum Gelingen von Unterricht beizutragen. Im Rahmen einer Zukunftswerkstatt hat das Leibniz-Netzwerk Unterrichtsforschung daher einen längerfristig angelegten Prozess gestartet, um sich mit dem Unterricht der Zukunft zu beschäftigen und zu versuchen, drängende Fragen und Handlungsfelder für die Unterrichtsqualitätsforschung zu identifizieren. Der vorliegende Beitrag stellt die Ergebnisse dieses initialen Austauschs dar. Zudem wird ein Einblick in die gegenwärtige Arbeit im Netzwerk gegeben, die aufbauend auf den Ergebnissen der Zukunftswerkstatt eine Weiterentwicklung der Unterrichtsqualitätsforschung anstrebt

HLA-DRB3/4/5 Matching Improves Outcome of Unrelated Hematopoietic Stem Cell Transplantation

The HLA-DRB3/4/5 loci are closely linked to the HLA-DRB1 gene. Mismatches in these loci occur with a frequency of about 8%–12% in otherwise 10/10 HLA-matched transplant pairs. There is preliminary evidence that these disparities may associate with increased acute graft-versus-host disease (GvHD) rates. The aim of this study was to analyze a large cohort of German patients and their donors for HLA-DRB3/4/5 compatibility and to correlate the HLA-DRB3/4/5 matching status with the outcome of unrelated hematopoietic stem cell transplantation (uHSCT). To this end, 3,410 patients and their respective donors were HLA-DRB3/4/5 and HLA-DPB1 typed by amplicon-based nextgeneration sequencing (NGS). All patients included received their first allogeneic transplant for malignant hematologic diseases between 2000 and 2014. Mismatches in the antigen recognition domain (ARD) of HLA-DRB3/4/5 genes were correlated with clinical outcome. HLA-DRB3/4/5 incompatibility was seen in 12.5% (n = 296) and 17.8% (n = 185) of the 10/10 and 9/10 HLA-matched cases, respectively. HLA-DRB3/4/5 mismatches in the ARD associated with a worse overall survival (OS), as shown in univariate (5-year OS: 46.1% vs. 39.8%, log-rank p = 0.038) and multivariate analyses [hazard ratio (HR) 1.25, 95% CI 1.02–1.54, p = 0.034] in the otherwise 10/10 HLAmatched subgroup. The worse outcome was mainly driven by a significantly higher nonrelapse mortality (HR 1.35, 95% CI 1.05–1.73, p = 0.017). In the 9/10 HLA-matched cases, the effect was not statistically significant. Our study results suggest that mismatches within the ARD of HLA-DRB3/4/5 genes significantly impact the outcome of otherwise fully matched uHSCT and support their consideration upon donor selection in the future

Transformative Materials to Create 3D Functional Human Tissue Models In Vitro in a Reproducible Manner

Recreating human tissues and organs in the petri dish to establish models as tools in biomedical sciences has gained momentum. These models can provide insight into mechanisms of human physiology, disease onset, and progression, and improve drug target validation, as well as the development of new medical therapeutics. Transformative materials play an important role in this evolution, as they can be programmed to direct cell behavior and fate by controlling the activity of bioactive molecules and material properties. Using nature as an inspiration, scientists are creating materials that incorporate specific biological processes observed during human organogenesis and tissue regeneration. This article presents the reader with state-of-the-art developments in the field of in vitro tissue engineering and the challenges related to the design, production, and translation of these transformative materials. Advances regarding (stem) cell sources, expansion, and differentiation, and how novel responsive materials, automated and large-scale fabrication processes, culture conditions, in situ monitoring systems, and computer simulations are required to create functional human tissue models that are relevant and efficient for drug discovery, are described. This paper illustrates how these different technologies need to converge to generate in vitro life-like human tissue models that provide a platform to answer health-based scientific questions.</p

Monitoring the electroactive cargo of extracellular vesicles can differentiate various cancer cell lines

Extracellular vesicles (EVs) are pivotal in cell-to-cell communication due to the array of cargo contained within these vesicles. EVs are considered important biomarkers for identification of disease, however most measurement approaches have focused on monitoring specific surface macromolecular targets. Our study focuses on exploring the electroactive component present within cargo from EVs obtained from various cancer and non-cancer cell lines using a disk carbon fiber microelectrode. Variations in the presence of oxidizable components were observed when the total cargo from EVs were measured, with the highest current detected in EVs from MCF7 cells. There were differences observed in the types of oxidizable species present within EVs from MCF7 and A549 cells. Single entity measurements showed clear spikes due to the detection of oxidizable cargo within EVs from MCF7 and A549 cells. These studies highlight the promise of monitoring EVs through the presence of varying electroactive components within the cargo and can drive a wave of new strategies towards specific detection of EVs for diagnosis and prognosis of various diseases

Gold marker displacement due to needle insertion during HDR-brachytherapy for treatment of prostate cancer: A prospective cone beam computed tomography and kilovoltage on-board imaging (kV-OBI) study

Abstract Purpose To evaluate gold marker displacement due to needle insertion during HDR-brachytherapy for therapy of prostate cancer. Patients and methods 18 patients entered into this prospective evaluation. Three gold markers were implanted into the prostate during the first HDR-brachytherapy procedure after the irradiation was administered. Three days after marker implantation all patients had a CT-scan for planning purpose of the percutaneous irradiation. Marker localization was defined on the digitally-reconstructed-radiographs (DRR) for daily (VMAT technique) or weekly (IMRT) set-up error correction. Percutaneous therapy started one week after first HDR-brachytherapy. After the second HDR-brachytherapy, two weeks after first HDR-brachtherapy, a cone-beam CT-scan was done to evaluate marker displacement due to needle insertion. In case of marker displacement, the actual positions of the gold markers were adjusted on the DRR. Results The value of the gold marker displacement due to the second HDR-brachytherapy was analyzed in all patients and for each gold marker by comparison of the marker positions in the prostate after soft tissue registration of the prostate of the CT-scans prior the first and second HDR-brachytherapy. The maximum deviation was 5 mm, 7 mm and 12 mm for the anterior-posterior, lateral and superior-inferior direction. At least one marker in each patient showed a significant displacement and therefore new marker positions were adjusted on the DRRs for the ongoing percutaneous therapy. Conclusions Needle insertion in the prostate due to HDR-brachytherapy can lead to gold marker displacements. Therefore, it is necessary to verify the actual position of markers after the second HDR-brachytherapy. In case of significant deviations, a new DRR with the adjusted marker positions should be generated for precise positioning during the ongoing percutaneous irradiation.</p