Recent Submissions

  • Journal Article

    Longtime Neurologic Outcome of Extracorporeal Membrane Oxygenation and Non Extracorporeal Membrane Oxygenation Acute Respiratory Distress Syndrome Survivors 

    Harnisch, Lars-Olav; Riech, Sebastian; Mueller, Marion; Gramueller, Vanessa; Quintel, Michael; Moerer, Onnen
    Journal of Clinical Medicine 2019; 8(7): Art. 1020
    Neurologic complications following acute respiratory distress syndrome (ARDS) are well described, however, information on the neurologic outcome regarding peripheral nervous system complications in critically ill ARDS patients, especially those who received extracorporeal membrane oxygenation (ECMO) are lacking. In this prospective observational study 28 ARDS patients who survived after ECMO or conventional nonECMO treatment were examined for neurological findings. Nine patients had findings related to cranial nerve innervation, which di ered between ECMO and nonECMO patients (p = 0.031). ECMO patients had severely increased patella tendon reflex (PTR) reflex levels (p = 0.027 vs. p = 0.125) as well as gastrocnemius tendon reflex (GTR) (p = 0.041 right, p = 0.149 left) were a ected on the right, but not on the left side presumably associated with ECMO cannulation. Paresis (14.3% of patients) was only found in the ECMO group (p = 0.067). Paresthesia was frequent (nonECMO 53.8%, ECMO 62.5%; p = 0.064), in nonECMO most frequently due to initial trauma and polyneuropathy, in the ECMO group mainly due to impairments of N. cutaneus femoris lateralis (4 vs. 0; p = 0.031). Besides well-known central neurologic complications, more subtle complications were detected by thorough clinical examination. These findings are su cient to hamper activities of daily living and impair quality of life and psychological health and are presumably directly related to ECMO therapy.
    View Document Abstract
  • Journal Article

    Changes in Musculoskeletal System and Metabolism in Osteoporotic Rats Treated With Urocortin 

    Saul, Dominik; Geisberg, Laura Katharina; Gehle, Torben; Hoffmann, Daniel Bernd; Tezval, Mohammad; Sehmisch, Stephan; Komrakova, Marina
    Frontiers in Endocrinology 2019; 10: Art. 400
    Objective: In aging population, postmenopausal osteoporosis and decline of musculoskeletal function, referred to as "frailty syndrome" lead to loss of bone and muscle, causing falls, and fall-related injuries. To limit the impact of this portentous duo, simultaneous treatment of both is needed. Urocortin (UCN) has been reported to improve osteoporotic bone properties while its effect on muscle has not been addressed yet. Design and Methods: We aimed to investigate the effect of urocortin in vivo on skeletal muscle structure in osteopenic rats. Sixty Sprague-Dawley rats were divided into five groups: four were ovariectomized (OVX) and one underwent sham operation (SHAM). One ovariectomized group was left untreated (OVX), while one was treated with urocortin s.c. in 3 μg/kg body weight (bw) (OVX+UCN low), one with 30 μg/kg (OVX+UCN high), while one group was treated with estradiol orally (OVX+E: 0.2 mg/kg bw), each for 35 days. Mm. gastrocnemius, longissimus, and soleus were isolated and capillary density as well as diameters of type I and II fibers were measured. In addition, we examined the effect of UCN on tibia using biomechanical, micro-CT and ashing analysis and investigated the blood serum. Results: We demonstrated a positive effect of UCN on M. soleus, in which fiber diameter was positively influenced. The biomechanical and structural parameters of bone were not changed in UCN treated rats. The higher cholesterol, glucose and triglyceride levels in the "UCN high" group raise concern about this treatment. Conclusions: Our results portray urocortin as a substance that can be assessed for future therapeutic treatments of estrogen deficiency. New and Noteworthy: Urocortin has a positive effect on M. soleus (diameter). Urocortin raises serum cholesterol and triglyceride levels. Bone tissue was not affected by UCN.
    View Document Abstract
  • Journal Article

    Growth hormone secretagogues: history, mechanism of action and clinical development 

    Ishida, Junichi; Saitoh, Masakazu; Ebner, Nicole; Springer, Jochen; Anker, Stefan D.; Haehling, Stephan von
    Journal of Cachexia, Sarcopenia and Muscle 2019; 2(2)
    Growth hormone secretagogues (GHSs) are a generic term to describe compounds which increase growth hormone (GH) release. GHSs include agonists of the growth hormone secretagogue receptor (GHS‐R), whose natural ligand is ghrelin, and agonists of the growth hormone‐releasing hormone receptor (GHRH‐R), to which the growth hormonereleasing hormone (GHRH) binds as a native ligand. Several GHSs have been developed with a view to treating or diagnosisg of GH deficiency, which causes growth retardation, gastrointestinal dysfunction and altered body composition, in parallel with extensive research to identify GHRH, GHS‐R and ghrelin. This review will focus on the research history and the pharmacology of each GHS, which reached randomized clinical trials. Furthermore, we will highlight the publicly disclosed clinical trials regarding GHSs.
    View Document Abstract
  • Journal Article

    Modulation of Conflict Processing by Theta-Range tACS over the Dorsolateral Prefrontal Cortex 

    Lehr, Albert; Henneberg, Niklas; Nigam, Tarana; Paulus, Walter; Antal, Andrea
    Neural Plasticity 2019; 2019 p.1-13: Art. 6747049
    Behavioral response conflict arises in the color-word Stroop task and triggers the cognitive control network. Midfrontal theta-band oscillations correlate with adaptive control mechanisms during and after conflict resolution. In order to prove causality, in two experiments, we applied transcranial alternating current stimulation (tACS) at 6 Hz to the dorsolateral prefrontal cortex (DLPFC) during Stroop task performance. Sham stimulation served as a control in both experiments; 9.7 Hz tACS served as a nonharmonic alpha band control in the second experiment. We employed generalized linear mixed models for analysis of behavioral data. Accuracy remained unchanged by any type of active stimulation. Over both experiments, the Stroop effect (response time difference between congruent and incongruent trials) was reduced by 6 Hz stimulation as compared to sham, mainly in trials without prior conflict adaptation. Alpha tACS did not modify the Stroop effect. Theta tACS can both reduce the Stroop effect and modulate adaptive mechanisms of the cognitive control network, suggesting midfrontal theta oscillations as causally involved in cognitive control.
    View Document Abstract
  • Journal Article

    Fluorinated nanobodies for targeted molecular imaging of biological samples using nanoscale secondary ion mass spectrometry 

    Kabatas, Selda; Agüi-Gonzalez, Paola; Hinrichs, Rena; Jähne, Sebastian; Opazo, Felipe; Diederichsen, Ulf; Rizzoli, Silvio O.; Phan, Nhu T. N.
    Journal of Analytical Atomic Spectrometry 2019; 34(6) p.1083-1087
    Molecular imaging of targeted large biomolecules has been restricted in SIMS due to the limited number of probes containing SIMS-detectable isotopes. We introduce here new 19F-containing molecules that can be conjugated in a site-specific manner to nanobodies able to recognize fluorescent proteins (FPs) or mouse immunoglobulins (Igs). In this work, we demonstrate that it is possible to use the 19F-nanobodies to reveal the location of several cellular proteins previously tagged with FPs or Igs. This enables specific bio-imaging in SIMS for a vast repertoire of biomolecules, offering new opportunities to study specific structural and functional molecular interactions in biological specimens.
    View Document Abstract
  • Journal Article

    Reorientational Dynamics of Amyloid-β from NMR Spin Relaxation and Molecular Simulation 

    Rezaei-Ghaleh, Nasrollah; Parigi, Giacomo; Zweckstetter, Markus
    The Journal of Physical Chemistry Letters 2019; 10(12) p.3369-3375
    Amyloid-β (Aβ) aggregation is a hallmark of Alzheimer's disease. As an intrinsically disordered protein, Aβ undergoes extensive dynamics on multiple length and time scales. Access to a comprehensive picture of the reorientational dynamics in Aβ requires therefore the combination of complementary techniques. Here, we integrate 15N spin relaxation rates at three magnetic fields with microseconds-long molecular dynamics simulation, ensemble-based hydrodynamic calculations, and previously published nanosecond fluorescence correlation spectroscopy to investigate the reorientational dynamics of Aβ1-40 (Aβ40) at single-residue resolution. The integrative analysis shows that librational and dihedral angle fluctuations occurring at fast and intermediate time scales are not sufficient to decorrelate orientational memory in Aβ40. Instead, slow segmental motions occurring at ∼5 ns are detected throughout the Aβ40 sequence and reach up to ∼10 ns for selected residues. We propose that the modulation of time scales of reorientational dynamics with respect to intra- and intermolecular diffusion plays an important role in disease-related Aβ aggregation.
    View Document Abstract
  • Journal Article

    Molecular phenotyping of the surfaceome of migratory chondroprogenitors and mesenchymal stem cells using biotinylation, glycocapture and quantitative LC-MS/MS proteomic analysis 

    Matta, Csaba; Boocock, David J.; Fellows, Christopher R.; Miosge, Nicolai; Dixon, James E.; Liddell, Susan; Smith, Julia; Mobasheri, Ali
    Scientific Reports 2019; 9(1): Art. 9018
    The complement of cell surface proteins, collectively referred to as the surfaceome, is a useful indicator of normal differentiation processes, and the development of pathologies such as osteoarthritis (OA). We employed biochemical and proteomic tools to explore the surfaceome and to define biomarkers in chondrogenic progenitor cells (CPC) derived from human OA knee articular cartilage. These cells have great therapeutic potential, but their unexplored biology limits their clinical application. We performed biotinylation combined with glycocapture and high throughput shotgun proteomics to define the surface proteome of human bone marrow mesenchymal stem cells (MSCs) and human CPCs. We prepared cell surface protein-enriched fractions from MSCs and CPCs, and then a proteomic approach was used to compare and evaluate protein changes between undifferentiated MSCs and CPCs. 1256 proteins were identified in the study, of which 791 (63%) were plasma membrane, cell surface or extracellular matrix proteins. Proteins constituting the surfaceome were annotated and categorized. Our results provide, for the first time, a repository of quantitative proteomic data on the surfaceome of two closely related cell types relevant to cartilage biology and OA. These results may provide novel insights into the transformation of the surfaceome during chondrogenic differentiation and phenotypic changes during OA development.
    View Document Abstract
  • Journal Article

    Ketogenic diet ameliorates axonal defects and promotes myelination in Pelizaeus–Merzbacher disease 

    Stumpf, Sina K.; Berghoff, Stefan A.; Trevisiol, Andrea; Spieth, Lena; Düking, Tim; Schneider, Lennart V.; Schlaphoff, Lennart; Dreha-Kulaczewski, Steffi; Bley, Annette; Burfeind, Dinah; et al.
    Kusch, KathrinMitkovski, MisoRuhwedel, TorbenGuder, PhilippRöhse, HeikoDenecke, JonasGärtner, JuttaMöbius, WiebkeNave, Klaus-ArminSaher, Gesine
    Acta Neuropathologica 2019; 138(1) p.147-161
    Pelizaeus-Merzbacher disease (PMD) is an untreatable and fatal leukodystrophy. In a model of PMD with perturbed blood-brain barrier integrity, cholesterol supplementation promotes myelin membrane growth. Here, we show that in contrast to the mouse model, dietary cholesterol in two PMD patients did not lead to a major advancement of hypomyelination, potentially because the intact blood-brain barrier precludes its entry into the CNS. We therefore turned to a PMD mouse model with preserved blood-brain barrier integrity and show that a high-fat/low-carbohydrate ketogenic diet restored oligodendrocyte integrity and increased CNS myelination. This dietary intervention also ameliorated axonal degeneration and normalized motor functions. Moreover, in a paradigm of adult remyelination, ketogenic diet facilitated repair and attenuated axon damage. We suggest that a therapy with lipids such as ketone bodies, that readily enter the brain, can circumvent the requirement of a disrupted blood-brain barrier in the treatment of myelin disease.
    View Document Abstract
  • Journal Article

    Walking pathways with positive feedback loops reveal DNA methylation biomarkers of colorectal cancer 

    Kel, Alexander; Boyarskikh, Ulyana; Stegmaier, Philip; Leskov, Leonid S.; Sokolov, Andrey V.; Yevshin, Ivan; Mandrik, Nikita; Stelmashenko, Daria; Koschmann, Jeannette; Kel-Margoulis, Olga; et al.
    Krull, MathiasMartínez-Cardús, AnnaMoran, SebastianEsteller, ManelKolpakov, FedorFilipenko, MaximWingender, Edgar
    BMC Bioinformatics 2019; 20(S4): Art. 119
    Background: The search for molecular biomarkers of early-onset colorectal cancer (CRC) is an important but still quite challenging and unsolved task. Detection of CpG methylation in human DNA obtained from blood or stool has been proposed as a promising approach to a noninvasive early diagnosis of CRC. Thousands of abnormally methylated CpG positions in CRC genomes are often located in non-coding parts of genes. Novel bioinformatic methods are thus urgently needed for multi-omics data analysis to reveal causative biomarkers with a potential driver role in early stages of cancer. Methods: We have developed a method for finding potential causal relationships between epigenetic changes (DNA methylations) in gene regulatory regions that affect transcription factor binding sites (TFBS) and gene expression changes. This method also considers the topology of the involved signal transduction pathways and searches for positive feedback loops that may cause the carcinogenic aberrations in gene expression. We call this method “Walking pathways”, since it searches for potential rewiring mechanisms in cancer pathways due to dynamic changes in the DNA methylation status of important gene regulatory regions (“epigenomic walking”). Results: In this paper, we analysed an extensive collection of full genome gene-expression data (RNA-seq) and DNA methylation data of genomic CpG islands (using Illumina methylation arrays) generated from a sample of tumor and normal gut epithelial tissues of 300 patients with colorectal cancer (at different stages of the disease) (data generated in the EU-supported SysCol project). Identification of potential epigenetic biomarkers of DNA methylation was performed using the fully automatic multi-omics analysis web service “My Genome Enhancer” (MGE) (my-genome-enhancer.com). MGE uses the database on gene regulation TRANSFAC®, the signal transduction pathways database TRANSPATH®, and software that employs AI (artificial intelligence) methods for the analysis of cancer-specific enhancers. (Continued from previous page) Conclusions: The identified biomarkers underwent experimental testing on an independent set of blood samples from patients with colorectal cancer. As a result, using advanced methods of statistics and machine learning, a minimum set of 6 biomarkers was selected, which together achieve the best cancer detection potential. The markers include hypermethylated positions in regulatory regions of the following genes: CALCA, ENO1, MYC, PDX1, TCF7, ZNF43.
    View Document Abstract
  • Journal Article

    Progesterone improves survival in hepatoma cachexia rat model 

    Suzuki, Tsuyoshi; Ebner, Nicole; Palus, Sandra; Haehling, Stephan von; Springer, Jochen
    Journal of Cachexia, Sarcopenia and Muscle
    Background: Medroxyprogesterone (MPA) and megestrol acetate (MA) are synthetic progesterone derivates. Progestagen is an approved drug for cancer cachexia in the USA and some Europe. These agents have been described to increase appetite and to lead to weight gain. However, the effects on survival are still unknown. The aim of this study was to evaluate the effects of progesterone on survival, cardiac function as well as appetite and body weight in the Yoshida hepatoma AH‐130 rat cancer cachexia model. Methods: In this study the effects of progesterone were tested in cachectic tumor bearing rats. Rats were treated with 0.5, 5 or 50mg/kg/d, respectively or placebo daily, starting one day after tumor inoculation for a period of 16 days. Cardiac function was analyzed by echocardiography at baseline and at day 11. Food intake was assessed before tumor inoculation and at day 11. Body weight and body composition were evaluated at the beginning and the end of study or day of euthanasia. Results: Survival was significantly improved by 5 mg/kg/d (HR: 0.48, 95%CI: 0.24‐0.95, p=0.0356). However, there was no significant difference between the progesterone treatment groups compared to placebo in body weight change and body composition, as well as food intake on day 11. Cardiac function also showed no significant difference compared to placebo. Conclusion: Progesterone improves survival, but has no beneficial effects on cardiac function, body weight and food intake in this aggressive hepatoma cancer cachexia rat model. Further studies are needed to elucidate the mechanism of the survival benefit.
    View Document Abstract
  • Journal Article

    Cytosolic Trapping of a Mitochondrial Heat Shock Protein Is an Early Pathological Event in Synucleinopathies 

    Szegő, Éva M.; Dominguez-Meijide, Antonio; Gerhardt, Ellen; König, Annekatrin; Koss, David J.; Li, Wen; Pinho, Raquel; Fahlbusch, Christiane; Johnson, Mary; Santos, Patricia; et al.
    Villar-Piqué, AnnaThom, TobiasRizzoli, SilvioSchmitz, MatthiasLi, JiayiZerr, IngaAttems, JohannesJahn, OlafOuteiro, Tiago F.
    Cell Reports 2019; 28(1) p.65-77
    Alpha-synuclein (aSyn) accumulates in intracellular inclusions in synucleinopathies, but the molecular mechanisms leading to disease are unclear. We identify the 10 kDa heat shock protein (HSP10) as a mediator of aSyn-induced mitochondrial impairments in striatal synaptosomes. We find an age-associated increase in the cytosolic levels of HSP10, and a concomitant decrease in the mitochondrial levels, in aSyn transgenic mice. The levels of superoxide dismutase 2, a client of the HSP10/HSP60 folding complex, and synaptosomal spare respiratory capacity are also reduced. Overexpression of HSP10 ameliorates aSyn-associated mitochondrial dysfunction and delays aSyn pathology in vitro and in vivo. Altogether, our data indicate that increased levels of aSyn induce mitochondrial deficits, at least partially, by sequestering HSP10 in the cytosol and preventing it from acting in mitochondria. Importantly, these alterations manifest first at presynaptic terminals. Our study not only provides mechanistic insight into synucleinopathies but opens new avenues for targeting underlying cellular pathologies.
    View Document Abstract
  • Journal Article

    Long-Term Impact of Toxoplasma gondii Infection on Human Monocytes 

    Ehmen, Hauke G.; Lüder, Carsten G. K.
    Frontiers in Cellular and Infection Microbiology 2019; 9: Art. 235
    Toxoplasma gondii is a prevalent parasite of mammals and birds including up to 30% of humans world-wide. Primary infection of immunocompetent hosts leads to a robust cell-mediated immune response, which controls but does not clear the infection, thus enabling long-term parasite persistence in brain and muscle tissues. Chronic toxoplasmosis in mice is associated with resistance to heterologous pathogens and this has been related to increased numbers of inflammatory monocytes. Here we have analyzed whether chronic T. gondii infection impacts the subset distribution and the phenotype of peripheral human monocytes in vivo and their responses to parasite infection in vitro. CD14+ monocytes from T. gondii-seropositive blood donors expressed significantly less FcγRIII (CD16) than those from seronegative controls, but they did not show a shift in the distribution of classical, intermediate and non-classical monocyte subpopulations. Percentages of CD62L+ and CD64+ monocytes were however decreased and increased, respectively, in chronically infected individuals as compared to naïve controls. Infection of monocyte-enriched PBMCs from both seropositive and seronegative individuals with T. gondii led to an increase of CD14+CD16− classical monocytes and a decrease of CD14+CD16+ double positive monocytes. Remarkably, after in vitro parasite infection, expression of the chemokine receptor CCR2 was severely impaired in monocytes from both, individuals with chronic toxoplasmosis and seronegative controls. In contrast, only monocytes from chronically infected humans but not those from controls dose-dependently up-regulated HLA-DR, DP, DQ expression following in vitro infection. Furthermore, monocyte-enriched PBMCs from seropositive individuals up-regulated IL-12 mRNA more vigorously after in vitro infection than cells from naïve controls. Collectively, our results establish that infection of humans with T. gondii exerts long-term effects on the phenotype and responsiveness of blood monocytes. This may have important implications for innate immune responses to T. gondii and unrelated pathogens.
    View Document Abstract
  • Journal Article

    Effectiveness of a universal adhesive for repair bonding to composite and amalgam 

    Kanzow, Philipp; Baxter, Steffi; Rizk, Marta; Wassmann, Torsten; Wiegand, Annette
    Journal of Oral Science 2019; 61(2) p.343-350
    The study aimed to compare the repair bond strength of aged composite and amalgam repaired with resin composite after various mechanical and adhesive surface treatments. Specimens were aged by thermal cycling (10,000 cycles, 5-55°C) and randomly subjected to one of three surface treatments: diamond bur abrasion, aluminum oxide air abrasion, or silica coating. Conventional bonding or a universal adhesive with incorporated silane was applied afterward (each n = 16) and resin composite was attached. In the control groups (each n = 16), resin composite was attached using one of the above adhesives without prior mechanical surface conditioning. After further thermal cycling, the shear bond strength (SBS) and failure modes were assessed. Statistical analyses were performed using ANOVA, Weibull statistics, two sample t-tests, and Chi2-test (P < 0.05). The SBS of the repaired amalgam was significantly lower than that of the composite and mechanical pretreatment significantly increased SBS. The universal adhesive significantly improved the SBS of the repaired amalgam compared to the conventional bonding agent and mechanical pretreatment increased the number of cohesive/mixed failures. Amalgam restorations may be repaired using resin composites, but the resulting SBS is lower than that obtained with composite
    View Document Abstract
  • Journal Article

    Inhibition of the Hypoxia-Inducible Factor 1α–Induced Cardiospecific HERNA1 Enhance-Templated RNA Protects From Heart Disease 

    Mirtschink, Peter; Bischof, Corinne; Pham, Minh-Duc; Sharma, Rahul; Khadayate, Sanjay; Rossi, Geetha; Fankhauser, Niklaus; Traub, Shuyang; Sossalla, Samuel; Hagag, Eman; et al.
    Berthonneche, CorinneSarre, AlexandreStehr, Sebastian. N.Grote, PhillipPedrazzini, ThierryDimmeler, StefanieKrek, WilhelmKrishnan, Jaya
    Circulation 2019; 139(24) p.2778-2792
    BACKGROUND: Enhancers are genomic regulatory elements conferring spatiotemporal and signal-dependent control of gene expression. Recent evidence suggests that enhancers can generate noncoding enhancer RNAs, but their (patho)biological functions remain largely elusive. METHODS: We performed chromatin immunoprecipitation-coupled sequencing of histone marks combined with RNA sequencing of left ventricular biopsies from experimental and genetic mouse models of human cardiac hypertrophy to identify transcripts revealing enhancer localization, conservation with the human genome, and hypoxia-inducible factor 1α dependence. The most promising candidate, hypoxia-inducible enhancer RNA ( HERNA)1, was further examined by investigating its capacity to modulate neighboring coding gene expression by binding to their gene promoters by using chromatin isolation by RNA purification and λN-BoxB tethering-based reporter assays. The role of HERNA1 and its neighboring genes for pathological stress-induced growth and contractile dysfunction, and the therapeutic potential of HERNA1 inhibition was studied in gapmer-mediated loss-of-function studies in vitro using human induced pluripotent stem cell-derived cardiomyocytes and various in vivo models of human pathological cardiac hypertrophy. RESULTS: HERNA1 is robustly induced on pathological stress. Production of HERNA1 is initiated by direct hypoxia-inducible factor 1α binding to a hypoxia-response element in the histoneH3-lysine27acetylation marks-enriched promoter of the enhancer and confers hypoxia responsiveness to nearby genes including synaptotagmin XVII, a member of the family of membrane-trafficking and Ca2+-sensing proteins and SMG1, encoding a phosphatidylinositol 3-kinase-related kinase. Consequently, a substrate of SMG1, ATP-dependent RNA helicase upframeshift 1, is hyperphoshorylated in a HERNA1- and SMG1-dependent manner. In vitro and in vivo inactivation of SMG1 and SYT17 revealed overlapping and distinct roles in modulating cardiac hypertrophy. Finally, in vivo administration of antisense oligonucleotides targeting HERNA1 protected mice from stress-induced pathological hypertrophy. The inhibition of HERNA1 postdisease development reversed left ventricular growth and dysfunction, resulting in increased overall survival. CONCLUSIONS: HERNA1 is a novel heart-specific noncoding RNA with key regulatory functions in modulating the growth, metabolic, and contractile gene program in disease, and reveals a molecular target amenable to therapeutic exploitation.
    View Document Abstract
  • Journal Article

    Robustness of testing procedures for confirmatory subpopulation analyses based on a continuous biomarker 

    Graf, Alexandra Christine; Wassmer, Gernot; Friede, Tim; Gera, Roland Gerard; Posch, Martin
    Statistical Methods in Medical Research 2018; 28(6) p.1879-1892
    With the advent of personalized medicine, clinical trials studying treatment effects in subpopulations are receiving increasing attention. The objectives of such studies are, besides demonstrating a treatment effect in the overall population, to identify subpopulations, based on biomarkers, where the treatment has a beneficial effect. Continuous biomarkers are often dichotomized using a threshold to define two subpopulations with low and high biomarker levels. If there is insufficient information on the dependence structure of the outcome on the biomarker, several thresholds may be investigated. The nested structure of such subpopulations is similar to the structure in group sequential trials. Therefore, it has been proposed to use the corresponding critical boundaries to test such nested subpopulations. We show that for biomarkers with a prognostic effect that is not adjusted for in the statistical model, the variability of the outcome may vary across subpopulations which may lead to an inflation of the family-wise type 1 error rate. Using simulations we quantify the potential inflation of testing procedures based on group sequential designs. Furthermore, alternative hypotheses tests that control the family-wise type 1 error rate under minimal assumptions are proposed. The methodological approaches are illustrated by a trial in depression.
    View Document Abstract
  • Journal Article

    Pten controls B‐cell responsiveness and germinal center reaction by regulating the expression of IgD BCR 

    Setz, Corinna S.; Khadour, Ahmad; Renna, Valerio; Iype, Joseena; Gentner, Eva; He, Xiaocui; Datta, Moumita; Young, Marc; Nitschke, Lars; Wienands, Jürgen; et al.
    Maity, Palash C.Reth, MichaelJumaa, Hassan
    The EMBO Journal 2019; 38(11): Art. e100249
    In contrast to other B-cell antigen receptor (BCR) classes, the function of IgD BCR on mature B cells remains largely elusive as mature B cells co-express IgM, which is sufficient for development, survival, and activation of B cells. Here, we show that IgD expression is regulated by the forkhead box transcription factor FoxO1, thereby shifting the responsiveness of mature B cells towards recognition of multivalent antigen. FoxO1 is repressed by phosphoinositide 3-kinase (PI3K) signaling and requires the lipid phosphatase Pten for its activation. Consequently, Pten-deficient B cells expressing knock-ins for BCR heavy and light chain genes are unable to upregulate IgD. Furthermore, in the presence of autoantigen, Pten-deficient B cells cannot eliminate the autoreactive BCR specificity by secondary light chain gene recombination. Instead, Pten-deficient B cells downregulate BCR expression and become unresponsive to further BCR-mediated stimulation. Notably, we observed a delayed germinal center (GC) reaction by IgD-deficient B cells after immunization with trinitrophenyl-ovalbumin (TNP-Ova), a commonly used antigen for T-cell-dependent antibody responses. Together, our data suggest that the activation of IgD expression by Pten/FoxO1 results in mature B cells that are selectively responsive to multivalent antigen and are capable of initiating rapid GC reactions and T-cell-dependent antibody responses.
    View Document Abstract
  • Journal Article

    Secretoneurin Is an Endogenous Calcium/Calmodulin-Dependent Protein Kinase II Inhibitor That Attenuates Ca 2+ -Dependent Arrhythmia 

    Ottesen, Anett H.; Carlson, Cathrine R.; Eken, Olav Søvik; Sadredini, Mani; Myhre, Peder L.; Shen, Xin; Dalhus, Bjørn; Laver, Derek R.; Lunde, Per Kristian; Kurola, Jouni; et al.
    Lunde, MarianneHoff, Jon ErikGodang, KristinSjaastad, IvarPettilä, VilleStridsberg, MatsLehnart, Stephan E.Edwards, Andrew G.Lunde, Ida G.Omland, TorbjørnStokke, Mathis K.Christensen, GeirRøsjø, HelgeLouch, William E.
    Circulation: Arrhythmia and Electrophysiology 2019; 12(4): Art. e007045
    Circulating SN (secretoneurin) concentrations are increased in patients with myocardial dysfunction and predict poor outcome. Because SN inhibits CaMKIIδ (Ca2+/calmodulin-dependent protein kinase IIδ) activity, we hypothesized that upregulation of SN in patients protects against cardiomyocyte mechanisms of arrhythmia. METHODS: Circulating levels of SN and other biomarkers were assessed in patients with catecholaminergic polymorphic ventricular tachycardia (CPVT; n=8) and in resuscitated patients after ventricular arrhythmia-induced cardiac arrest (n=155). In vivo effects of SN were investigated in CPVT mice (RyR2 [ryanodine receptor 2]-R2474S) using adeno-associated virus-9-induced overexpression. Interactions between SN and CaMKIIδ were mapped using pull-down experiments, mutagenesis, ELISA, and structural homology modeling. Ex vivo actions were tested in Langendorff hearts and effects on Ca2+ homeostasis examined by fluorescence (fluo-4) and patch-clamp recordings in isolated cardiomyocytes. RESULTS: SN levels were elevated in patients with CPVT and following ventricular arrhythmia-induced cardiac arrest. In contrast to NT-proBNP (N-terminal pro-B-type natriuretic peptide) and hs-TnT (high-sensitivity troponin T), circulating SN levels declined after resuscitation, as the risk of a new arrhythmia waned. Myocardial pro-SN expression was also increased in CPVT mice, and further adeno-associated virus-9-induced overexpression of SN attenuated arrhythmic induction during stress testing with isoproterenol. Mechanistic studies mapped SN binding to the substrate binding site in the catalytic region of CaMKIIδ. Accordingly, SN attenuated isoproterenol induced autophosphorylation of Thr287-CaMKIIδ in Langendorff hearts and inhibited CaMKIIδ-dependent RyR phosphorylation. In line with CaMKIIδ and RyR inhibition, SN treatment decreased Ca2+ spark frequency and dimensions in cardiomyocytes during isoproterenol challenge, and reduced the incidence of Ca2+ waves, delayed afterdepolarizations, and spontaneous action potentials. SN treatment also lowered the incidence of early afterdepolarizations during isoproterenol; an effect paralleled by reduced magnitude of L-type Ca2+ current. CONCLUSIONS: SN production is upregulated in conditions with cardiomyocyte Ca2+ dysregulation and offers compensatory protection against cardiomyocyte mechanisms of arrhythmia, which may underlie its putative use as a biomarker in at-risk patients.
    View Document Abstract
  • Journal Article

    Prevalence of post-concussion-like symptoms in the general population in Italy, The Netherlands and the United Kingdom 

    Voormolen, Daphne C.; Cnossen, Maryse C.; Polinder, Suzanne; Gravesteijn, Benjamin Y.; Von Steinbuechel, Nicole; Real, Ruben G.L.; Haagsma, Juanita A.
    Brain Injury p.1-9
    OBJECTIVES: To evaluate the frequency of post-concussion symptoms and prevalence and risk factors of post-concussion syndrome (PCS) in the general population, investigate the association between the Rivermead Post-Concussion Symptoms Questionnaire (RPQ) and self-perceived health, and evaluate differences between three European countries. METHODS: A web-based survey including the RPQ and EQ-5D was conducted among representative samples in three European countries. RESULTS: A total of 11,759 respondents completed the questionnaire. The most frequently reported symptom was fatigue (49.9%). Almost half (45.1%) of the respondents were classified as having PCS considering rating score 2 (three RPQ items with score ≥ 2) as a cut-off. Chronic health complaints were found as a significant risk factor for PCS. All items of the RPQ were positively correlated with the EQ-5D and the strongest positive correlation (0.633, p<0.001) was between RPQ item 'feeling depressed or tearful' and EQ-5D domain 'anxiety/depression'. CONCLUSIONS: We found a high frequency of post-concussion-like symptoms and PCS in the general population, indicating that these symptoms are not specific for patients with traumatic brain injury (TBI), and PCS is not a unique syndrome after TBI. Therefore, the use of post-concussion symptoms and PCS as outcome following mild TBI should be interpreted with caution.
    View Document Abstract
  • Journal Article

    Broad range of missense error frequencies in cellular proteins 

    Garofalo, Raffaella; Wohlgemuth, Ingo; Pearson, Michael; Lenz, Christof; Urlaub, Henning; Rodnina, Marina V.
    Nucleic Acids Research 2019; 47(6) p.2932-2945
    Assessment of the fidelity of gene expression is crucial to understand cell homeostasis. Here we present a highly sensitive method for the systematic Quantification of Rare Amino acid Substitutions (QRAS) using absolute quantification by targeted mass spectrometry after chromatographic enrichment of peptides with missense amino acid substitutions. By analyzing incorporation of near- and non-cognate amino acids in a model protein EF-Tu, we show that most of missense errors are too rare to detect by conventional methods, such as DDA, and are estimated to be between <10-7-10-5 by QRAS. We also observe error hotspots of up to 10-3 for some types of mismatches, including the G-U mismatch. The error frequency depends on the expression level of EF-Tu and, surprisingly, the amino acid position in the protein. QRAS is not restricted to any particular miscoding event, organism, strain or model protein and is a reliable tool to analyze very rare proteogenomic events.
    View Document Abstract
  • Journal Article

    Proteome Profiling by Label‐Free Mass Spectrometry Reveals Differentiated Response of Campylobacter jejuni 81–176 to Sublethal Concentrations of Bile Acids 

    Masanta, Wycliffe O.; Zautner, Andreas E.; Lugert, Raimond; Bohne, Wolfgang; Gross, Uwe; Leha, Andreas; Dakna, Mohammed; Lenz, Christof
    PROTEOMICS – Clinical Applications 2018; 13(3): Art. 1800083
    PURPOSE: Bile acids are crucial components of the intestinal antimicrobial defense and represent a significant stress factor for enteric pathogens. Adaptation processes of Campylobacter jejuni to this hostile environment are analyzed in this study by a proteomic approach. EXPERIMENTAL DESIGN: Proteome profiling by label-free mass spectrometry (SWATH-MS) has been used to characterize the adaptation of C. jejuni to sublethal concentrations of seven bile acids. RESULTS: The bile acids with the lowest inhibitory concentration (IC50 ), deoxycholic and chenodeoxycholic acid, induce the most significant proteome changes. Overall a downregulation of all basic biosynthetic pathways and a general decrease in the transcription machinery are found. Concurrently, an induction of factors involved in detoxification of reactive oxygen species, protein folding, and bile acid exporting efflux pumps is detected. Exposure to deoxycholic and chenodeoxycholic acid results in an increased expression of components of the more energy-efficient aerobic respiration pathway, while the anaerobic branches of the electron transport chain are down-expressed. CONCLUSIONS AND CLINICAL RELEVANCE: The results show that C. jejuni has a differentiated system of adaptation to bile acid stresses. The findings enhance the understanding of the pathogenesis of campylobacteriosis, especially for survival of C. jejuni in the human intestine, and may provide clues to future medical treatment.
    View Document Abstract

View more