Impairment of the neurotrophic signaling hub B-Raf contributes to motoneuron degeneration in spinal muscular atrophy.

Spinal muscular atrophy (SMA) is a motoneuron disease caused by deletions of the Survival of Motoneuron 1 gene (SMN1) and low SMN protein levels. SMN restoration is the concept behind a number of recently approved drugs which result in impressive yet limited effects. Since SMN has already been enhanced in treated patients, complementary SMN-independent approaches are needed. Previously, a number of altered signaling pathways which regulate motoneuron degeneration have been identified as candidate targets. However, signaling pathways form networks, and their connectivity is still unknown in SMA. Here, we used presymptomatic SMA mice to elucidate the network of altered signaling in SMA. The SMA network is structured in two clusters with AKT and 14-3-3 ζ/δ in their centers. Both clusters are connected by B-Raf as a major signaling hub. The direct interaction of B-Raf with 14-3-3 ζ/δ is important for an efficient neurotrophic activation of the MEK/ERK pathway and crucial for motoneuron survival. Further analyses in SMA mice revealed that both proteins were down-regulated in motoneurons and the spinal cord with B-Raf being reduced at presymptomatic stages. Primary fibroblasts and iPSC-derived motoneurons from SMA patients both showed the same pattern of down-regulation. This mechanism is conserved across species since a Caenorhabditis elegans SMA model showed less expression of the B-Raf homolog lin-45 Accordingly, motoneuron survival was rescued by a cell autonomous lin-45 expression in a C. elegans SMA model resulting in improved motor functions. This rescue was effective even after the onset of motoneuron degeneration and mediated by the MEK/ERK pathway.

ADHD in older adults - a scoping review.

OBJECTIVES: This scoping review presents an overview of the available research on symptoms, comorbidities, and associated challenges among older adults with ADHD. METHOD: The literature study followed Arksey and O'Malley's five-stage framework. The search was conducted in ProQuest Central, Scopus, PsycInfo, CINAHL, and PubMed. Articles were included if they were peer-reviewed, ethically approved primary studies, written in the English language, concerning ADHD, and including people 60 years of age and older. RESULTS: The review included 17 articles. Symptoms of ADHD persist throughout life. Older adults with ADHD experience similar core symptoms as younger individuals, but their manifestation and intensity may differ. The most common comorbidity found was mental illness, for example depression and anxiety. ADHD in older adults is linked to several challenges, such as difficulty with relationships and social isolation. CONCLUSION: Older adults with ADHD face various symptoms, comorbidities, and challenges that affect their quality of life. Age-related changes can amplify ADHD symptoms and increase the perceived burden of illness. More research is needed to understand the complex relationship between these factors and enable tailored interventions to improve their quality of life and well-being.

Interdisciplinary studies on Coxiella burnetii: From molecular to cellular, to host, to one health research.

The Q-GAPS (Q fever GermAn interdisciplinary Program for reSearch) consortium was launched in 2017 as a German consortium of more than 20 scientists with exceptional expertise, competence, and substantial knowledge in the field of the Q fever pathogen Coxiella (C.) burnetii. C. burnetii exemplifies as a zoonotic pathogen the challenges of zoonotic disease control and prophylaxis in human, animal, and environmental settings in a One Health approach. An interdisciplinary approach to studying the pathogen is essential to address unresolved questions about the epidemiology, immunology, pathogenesis, surveillance, and control of C. burnetii. In more than five years, Q-GAPS has provided new insights into pathogenicity and interaction with host defense mechanisms. The consortium has also investigated vaccine efficacy and application in animal reservoirs and identified expanded phenotypic and genotypic characteristics of C. burnetii and their epidemiological significance. In addition, conceptual principles for controlling, surveilling, and preventing zoonotic Q fever infections were developed and prepared for specific target groups. All findings have been continuously integrated into a Web-based, interactive, freely accessible knowledge and information platform (www.q-gaps.de), which also contains Q fever guidelines to support public health institutions in controlling and preventing Q fever. In this review, we will summarize our results and show an example of how an interdisciplinary consortium provides knowledge and better tools to control a zoonotic pathogen at the national level.

Drugging an undruggable pocket on KRAS.

The 3 human RAS genes, KRAS, NRAS, and HRAS, encode 4 different RAS proteins which belong to the protein family of small GTPases that function as binary molecular switches involved in cell signaling. Activating mutations in RAS are among the most common oncogenic drivers in human cancers, with KRAS being the most frequently mutated oncogene. Although KRAS is an excellent drug discovery target for many cancers, and despite decades of research, no therapeutic agent directly targeting RAS has been clinically approved. Using structure-based drug design, we have discovered BI-2852 (1), a KRAS inhibitor that binds with nanomolar affinity to a pocket, thus far perceived to be "undruggable," between switch I and II on RAS; 1 is mechanistically distinct from covalent KRASG12C inhibitors because it binds to a different pocket present in both the active and inactive forms of KRAS. In doing so, it blocks all GEF, GAP, and effector interactions with KRAS, leading to inhibition of downstream signaling and an antiproliferative effect in the low micromolar range in KRAS mutant cells. These findings clearly demonstrate that this so-called switch I/II pocket is indeed druggable and provide the scientific community with a chemical probe that simultaneously targets the active and inactive forms of KRAS.

Creativity is associated with higher well-being and more positive COVID-19 experience.

We investigated whether creativity is associated with higher well-being and more positive COVID-19 experience. Participants (N = 252) filled out a creativity measure during the COVID-19 pandemic, they rated their positive affect and stress experience in the last month, their satisfaction with life, and indicated the extent to which they perceived COVID-19 as a positive experience. More creative individuals were more satisfied with their lives after controlling for perceived stress and personality. Results of a serial mediation showed that creativity fostered more positive emotions, which lowered perceived stress, which then led to a more positive COVID-19 experience. Findings add to the literature showing the beneficial effects of creativity on well-being, and point to the utility of introducing interventions that would promote creative thinking to improve quality of life and resilience to life adversities.

Comparison of Different Commercially Available Enzyme-Linked Immunosorbent Assays with Immunofluorescence Test for Detection of Phase II IgG and IgM Antibodies to Coxiella burnetii.

Several commercially available enzyme-linked immunosorbent assays (ELISAs) for the detection of phase II IgG or IgM antibodies against Coxiella burnetii were compared. In addition, an indirect immunofluorescence test was used as a confirmation test. In all, 70 serum samples for IgG and 43 serum samples for IgM were tested. The ELISAs showed large differences in sensitivity and specificity, which led to a partially high ratio of false-negative determinations. The most convincing test was PanBio from Abbott, which unfortunately can only test IgG but not IgM.

Metalloprotease-mediated cleavage of PlexinD1 and its sequestration to actin rods in the motoneuron disease spinal muscular atrophy (SMA).

Cytoskeletal rearrangement during axon growth is mediated by guidance receptors and their ligands which act either as repellent, attractant or both. Regulation of the actin cytoskeleton is disturbed in Spinal Muscular Atrophy (SMA), a devastating neurodegenerative disease affecting mainly motoneurons, but receptor-ligand interactions leading to the dysregulation causing SMA are poorly understood. In this study, we analysed the role of the guidance receptor PlexinD1 in SMA pathogenesis. We showed that PlexinD1 is cleaved by metalloproteases in SMA and that this cleavage switches its function from an attractant to repellent. Moreover, we found that the PlexinD1 cleavage product binds to actin rods, pathological aggregate-like structures which had so far been described for age-related neurodegenerative diseases. Our data suggest a novel disease mechanism for SMA involving formation of actin rods as a molecular sink for a cleaved PlexinD1 fragment leading to dysregulation of receptor signaling.

Serological survey of Bartonella spp., Borrelia burgdorferi, Brucella spp., Coxiella burnetii, Francisella tularensis, Leptospira spp., Echinococcus, Hanta-, TBE- and XMR-virus infection in employees of two forestry enterprises in North Rhine-Westphalia, Germany, 2011-2013.

We initiated a survey to collect basic data on the frequency and regional distribution of various zoonoses in 722 employees of forestry enterprises in the German state of North Rhine-Westphalia (NRW) from 2011 to 2013. Exposures associated with seropositivity were identified to give insight into the possible risk factors for infection with each pathogen. 41.2% of participants were found to be seropositive for anti-Bartonella IgG, 30.6% for anti-Borrelia burgdorferi IgG, 14.2% for anti-Leptospira IgG, 6.5% for anti-Coxiella burnetii IgG, 6.0% for anti-Hantavirus IgG, 4.0% for anti-Francisella tularensis IgG, 3.4% for anti-TBE-virus IgG, 1.7% for anti-Echinococcus IgG, 0.0% for anti-Brucella IgG and anti-XMRV IgG. Participants seropositive for B. burgdorferi were 3.96 times more likely to be professional forestry workers (univariable analysis: OR 3.96; 95% CI 2.60-6.04; p<0.001); and participants seropositive for Hantavirus 3.72 times more likely (univariable analysis: OR 3.72; 95% CI 1.44-9.57; p=0.007). This study found a surprisingly high percentage of participants seropositive for anti-B. henselae IgG and for anti-F. tularensis IgG. The relatively high seroprevalence for anti-Leptospira IgG seen in this study could be related to living conditions rather than to exposure at work. No specific risk for exposure to C. burnetii and Echinococcus was identified, indicating that neither forestry workers nor office workers represent a risk population and that NRW is not a typical endemic area. Forestry workers appear to have higher risk for contact with B. burgdorferi-infected ticks and a regionally diverse risk for acquiring Hantavirus-infection. The regional epidemiology of zoonoses is without question of great importance for public health. Knowledge of the regional risk factors facilitates the development of efficient prevention strategies and the implementation of such prevention measures in a sustainable manner.

Regulation of memory B-cell survival by the BH3-only protein Puma.

Apoptosis is crucial for immune system homeostasis, including selection and survival of long-lived antibody-forming cells and memory cells. The interactions between proapoptotic and pro-survival proteins of the Bcl-2 family are critical for this process. In this report, we show that expression of the proapoptotic BH3-only Bcl-2 family member Puma was selectively up-regulated on in vitro activation with antigens or mitogens of both human and mouse B cells. Puma expression coincided in vivo, with the prosurvival Bcl-2 family member Mcl-1 within the germinal centers and its expression correlates with the germinal center like phenotype of Burkitt lymphoma. Experiments performed in Puma-deficient mice revealed that Puma is essential for apoptosis of mitogen-activated B cells in vitro and for the control of memory B-cell survival. In conclusion, using both human and murine models, our data show that Puma has a major role in the T cell- dependent B-cell immune response. These data demonstrate that Puma is a major regulator of memory B lymphocyte survival and therefore a key molecule in the control of the immune response.

Contemporary diagnostics for medically relevant fastidious microorganisms belonging to the genera Anaplasma,Bartonella,Coxiella,OrientiaandRickettsia.

Many of the human infectious pathogens-especially the zoonotic or vector-borne bacteria-are fastidious organisms that are difficult to cultivate because of their strong adaption to the infected host culminating in their near-complete physiological dependence on this environment. These bacterial species exhibit reduced multiplication rates once they are removed from their optimal ecological niche. This fact complicates the laboratory diagnosis of the disease and hinders the detection and further characterization of the underlying organisms, e.g. at the level of their resistance to antibiotics due to their slow growth. Here, we describe the current state of microbiological diagnostics for five genera of human pathogens with a fastidious laboratory lifestyle. For Anaplasma spp., Bartonella spp., Coxiella burnetii, Orientia spp. and Rickettsia spp., we will summarize the existing diagnostic protocols, the specific limitations for implementation of novel diagnostic approaches and the need for further optimization or expansion of the diagnostic armamentarium. We will reflect upon the diagnostic opportunities provided by new technologies including mass spectrometry and next-generation nucleic acid sequencing. Finally, we will review the (im)possibilities of rapidly developing new in vitro diagnostic tools for diseases of which the causative agents are fastidiously growing and therefore hard to detect.

BH3-only protein Puma contributes to death of antigen-specific T cells during shutdown of an immune response to acute viral infection.

During acute T cell immune responses to viral infection, antigen-specific T cells first proliferate and differentiate into effector cells, but after pathogen clearance most are deleted by apoptosis. The developmentally programmed death of antigen-specific T cells during shutdown of a T cell response is mediated by the Bcl-2-regulated apoptotic pathway and partly depends on the proapoptotic BH3-only protein Bim. However, loss of Bim enhanced survival of antigen-activated T cells to a lesser extent than Bcl-2 overexpression, indicating that other proapoptotic factors must contribute to T cell killing. In this study, we investigated the contributions of several BH3-only proteins to the shutdown of an acute T cell immune response in vivo. After infection with human herpes simplex virus (HSV-1), mice lacking Noxa, Bid, or Bad had a normal increase and subsequent decline in the numbers of antigen-specific CD8(+) T cells. In contrast, Puma-deficient mice showed an abnormally prolonged persistence of antigen-specific CD8(+) T cells in the spleen, associated with enhanced in vitro survival of these cells in the absence of cytokines. Puma was dispensable for viral clearance and also did not play a role in proliferation or activation of HSV-1-specific CD8(+) T cells in vivo. Collectively, these findings show that Puma contributes to the death of antigen-specific T cells during shutdown of an immune response.

Development of High-Throughput Multiplex Serology to Detect Serum Antibodies against Coxiella burnetii.

The causative agent of Q fever, the bacterium Coxiella burnetii (C. burnetii), has gained increasing interest due to outbreak events and reports about it being a potential risk factor for the development of lymphomas. In order to conduct large-scale studies for population monitoring and to investigate possible associations more closely, accurate and cost-effective high-throughput assays are highly desired. To address this need, nine C. burnetii proteins were expressed as recombinant antigens for multiplex serology. This technique enables the quantitative high-throughput detection of antibodies to multiple antigens simultaneously in a single reaction. Based on a reference group of 76 seropositive and 91 seronegative sera, three antigens were able to detect C. burnetii infections. Com1, GroEL, and DnaK achieved specificities of 93%, 69%, and 77% and sensitivities of 64%, 72%, and 47%, respectively. Double positivity to Com1 and GroEL led to a combined specificity of 90% and a sensitivity of 71%. In a subgroup of seropositives with an increased risk for chronic Q fever, the double positivity to these markers reached a specificity of 90% and a sensitivity of 86%. Multiplex serology enables the detection of antibodies against C. burnetii and appears well-suited to investigate associations between C. burnetii infections and the clinical manifestations in large-scale studies.

Concept of an Active Surveillance System for Q Fever in German Small Ruminants-Conflicts Between Best Practices and Feasibility.

Q fever is a zoonotic disease caused by the bacterium Coxiella burnetii. Inhalation of contaminated dust particles or aerosols originating from animals (esp. small ruminants) is the main source of human infection. Hence, an active early warning system for Q fever in German small ruminant livestock was conceptualized to prevent human infections. First, we describe the best practice for establishing this system before evaluating its feasibility, as the combination of both evokes conflicts. Vaginal swabs from all husbandry systems with a focus on reproductive females should pooled and investigated by PCR to detect C. burnetii-shedding animals. Multistage risk-based sampling shall be carried out at the flock level and within-flock level. At the flock level, all flocks that are at risk to transmit the pathogen to the public must be sampled. At the within-flock level, all primi- and multiparous females after lambing must be tested in order to increase the probability of identifying a positive herd. Sampling should be performed during the main lambing period and before migration in residential areas. Furthermore, individual animals should be tested before migration or exhibition to ensure a negative status. If a flock tests positive in at least one individual sample, then flock-specific preventive measures should be implemented. This approach implies huge financial costs (sample testing, action/control measures). Hence, taking the step to develop more feasible and affordable preventive measures, e.g., vaccinating small ruminant flocks, should replace testing wherever justifiable.

Chlamydia inhibit host cell apoptosis by degradation of proapoptotic BH3-only proteins.

Chlamydia are obligate intracellular bacteria that replicate in a vacuole inside a host cell. Chlamydial infection has been shown to protect the host cell against apoptotic stimuli. This is likely important for the ability of Chlamydia to reproduce in human cells. Here we show that resistance to apoptosis is conveyed by the destruction of the proapoptotic BH3-only proteins Bim/Bod, Puma, and Bad during infection. Apoptotic stimuli were blocked upstream of the mitochondrial activation of Bax/Bak. During infection with both species, Chlamydia trachomatis and Chlamydia pneumoniae, Bim protein gradually disappeared without noticeable changes in Bim mRNA. The disappearance was blocked by inhibitors of the proteasome. Infected cells retained sensitivity to Bim expressed by transfection, indicating functional relevance of the Bim disappearance. Fusion to Bim targeted the green fluorescent protein for destruction during infection. Analysis of truncation mutants showed that a short region of Bim containing the BH3 domain was sufficient for destruction during chlamydial infection. Like Bim, Puma and Bad proteins disappeared during infection. These results reveal a novel way by which microbes can interfere with the host cell's apoptotic machinery, and provide a molecular explanation of the cellular resistance to apoptosis during infection with Chlamydia.

Narrative Potential of Picture-Book Apps: A Media- and Interaction-Oriented Study.

Digital literature is playing an increasingly important role in children's everyday lives and opening up new paths for family literacy and early childhood education. However, despite positive effects of electronic books and picture-book apps on vocabulary learning, early writing, or phonological awareness, research findings on early narrative skills are ambiguous. Particularly, there still is a research gap regarding how app materiality affects children's story understanding. Thus, based on the ViSAR model for picture-book app analysis and data stemming from 12 digital reading dyads containing German monolingual 2- to 3-year-olds and their caregivers this study assessed the narrative potential of a commercial picture-book app and how this is used in interaction. Results of the media analysis showed that the app provides a high number of narrative animations. These animations could be used interactively to engage the child in the story. However, results of the interaction analysis showed that adult readers do not exploit this potential due to their strong concentration on operative prompts and instructions. Furthermore, an explorative analysis of the relation between adults' utterances and children's story comprehension provided preliminary indicators regarding how the length of reading duration and the number of utterances might relate to children's understanding of the story. Findings and methodological limitations of the study are discussed and combined didactically with practical recommendations on how to use narrative animations in interaction effectively.

[Hyperbaric oxygenation: utility in intensive therapy - part 2]. / Hyperbare Oxygenation - Stellenwert in der Intensivtherapie - Teil 2.

Gangrene, non-clostridial myonecroses and intracranial abscesses are clinical entities for which hyperbaric oxygenation can be used therapeutically. Mortality and invalidity can be reduced by this means. Except for gangrene, HBO is used merely as an adjuvant for these diseases when conventional surgical, antibiotic and intensive therapy measures are not sufficient. The action of HBO is based on several points of attack: it reduces the formation of oedema, inhibits the production of alpha-toxins of the Clostridia, has a bacteriotoxic action, increases the effectivity of antibiotics and improves the immune defense system. An early start is decisive for the success of HBO therapy.

[Hyperbaric Oxygenation - utility in intensive therapy - part 1]. / Hyperbare Oxygenation - Stellenwert in der Intensivtherapie - Teil 1.

Hyperbaric Oxygenation (HBO) is a medical treatment, in which a patient breathes 100 % oxygen inside a treatment chamber at a pressure higher than at sea level. The indication for HBO has to be considered whenever gas bubbles present in tissue or blood circulation (decompression sickness, gas embolism) or when a high oxygen delivery is of crucial importance (CO-intoxication, selected anaerobic soft tissue infections). HBO must never be considered as a substitute for conventional surgical or intensive care therapy, but as an adjuvant of (potentially) crucial importance.

Induction of apoptosis by Legionella pneumophila in mammalian cells requires the mitochondrial pathway for caspase activation.

Legionella pneumophila, the agent of human Legionnaire's disease is a Gram-negative, rod-shaped bacterium. During infection, the bacteria invade human cells and replicate intracellularly. L. pneumophila can induce apoptosis in human myeloid and epitheloid cells and this may contribute to the development of pathology and disease. However, the molecular mechanism of apoptosis induction is still uncertain. Here we investigate this process. Legionella efficiently induced apoptosis in myeloid cells, T cells and fibroblasts. Induction of apoptosis involved activation of the initiator caspase-9 and effector caspases. Caspase activity was required for cell death. Analysis of mutant cells showed that the death receptor pathway was not involved in Legionella-induced apoptosis. Surprisingly, caspase activity was found almost exclusively in cells that did not harbor bacteria. Infection with Legionella caused the activation of the pro-apoptotic protein Bax and the release of cytochrome c. Mouse embryonic fibroblasts deficient for Bax and/or Bak were protected from Legionella-induced caspase activation. These results show a clear contribution of the mitochondrial pathway to Legionella-induced apoptosis.

Direct inhibition of cytochrome c-induced caspase activation in vitro by Toxoplasma gondii reveals novel mechanisms of interference with host cell apoptosis.

The intracellular parasite Toxoplasma gondii is known to inhibit apoptosis of its host cell. The molecular mechanisms of this interference are, however, not yet completely understood. We show here that viable parasites prominently inhibited the activation of caspase 3/7 induced by cytochrome c, dATP and dithiothreitol in cytosolic extracts of human-derived Jurkat leukemic T cells. In contrast, granzyme B-induced caspase activity was only slightly diminished. De novo protein biosynthesis by T. gondii was dispensable for the inhibition of cytochrome c-induced caspase activation. Furthermore, a complete parasite lysate or, more importantly, molecules released by extracellular parasites mediated the interaction with the caspase cascade. The cell-free system applied here is thus a valuable tool to study the interaction of T. gondii and possibly other intracellular pathogens with host cell apoptosis.