Classification of neurons in the primate reticular formation and changes after recovery from pyramidal tract lesion

The reticular formation is important in primate motor control, both in health and during recovery after brain damage. Little is known about the different neurons present in the reticular nuclei. Here we recorded extracellular spikes from the reticular formation in five healthy female awake behaving monkeys (193 cells), and in two female monkeys 1 year after recovery from a unilateral pyramidal tract lesion (125 cells). Analysis of spike shape and four measures derived from the interspike interval distribution identified four clusters of neurons in control animals. Cluster 1 cellshadaslowfiringrate. Cluster 2 cell shad narrow spikes and irregular firing, which of ten included high-frequencybursts. Cluster3cellswere highly rhythmic and fast firing. Cluster 4 cells showed negative spikes. A separate population of 42 cells was antidromically identified as reticulospinal neurons in five anesthetized female monkeys. The distribution of spike width in these cells closely overlaid the distribution for cluster 2, leading us tentatively to suggest that cluster 2 included neurons with reticulospinal projections. In animals after corticospinal lesion, cells could be identified in all four clusters. The firing rate of cells in clusters 1 and 2 was increased in lesioned animals relative to control animals (by 52% and 60%, respectively); cells in cluster 2 were also more regular and more bursting in the lesioned animals. We suggest that changes in both membrane properties and local circuits within the reticular formation occur following lesioning, potentially increasing reticulospinal output to help compensate for lost corticospinal descending drive

Mycobacterium tuberculosis progresses through two phases of latent infection in humans

Little is known about the physiology of latent Mycobacterium tuberculosis infection. We studied the mutational rates of 24 index tuberculosis (TB) cases and their latently infected household contacts who developed active TB up to 5.25 years later, as an indication of bacterial physiological state and possible generation times during latent TB infection in humans. Here we report that the rate of new mutations in the M. tuberculosis genome decline dramatically after two years of latent infection (two-sided p < 0.001, assuming an 18 h generation time equal to log phase M. tuberculosis, with latency period modeled as a continuous variable). Alternatively, assuming a fixed mutation rate, the generation time increases over the latency duration. Mutations indicative of oxidative stress do not increase with increasing latency duration suggesting a lack of host or bacterial derived mutational stress. These results suggest that M. tuberculosis enters a quiescent state during latency, decreasing the risk for mutational drug resistance and increasing generation time, but potentially increasing bacterial tolerance to drugs that target actively growing bacteria.publishersversionpublishe

The Role of Immunological Synapse in Predicting the Efficacy of Chimeric Antigen Receptor (CAR) Immunotherapy

Chimeric Antigen Receptor (CAR) immunotherapy utilizes genetically-engineered immune cells that express a unique cell surface receptor that combines tumor antigen specificity with immune cell activation. In recent clinical trials, the adoptive transfer of CAR-modified immune cells (including CAR-T and CAR-NK cells) into patients has been remarkably successful in treating multiple refractory blood cancers. To improve safety and efficacy, and expand potential applicability to other cancer types, CARs with different target specificities and sequence modifications are being developed and tested by many laboratories. Despite the overall progress in CAR immunotherapy, conventional tools to design and evaluate the efficacy and safety of CAR immunotherapies can be inaccurate, time-consuming, costly, and labor-intensive. Furthermore, existing tools cannot always determine how responsive individual patients will be to a particular CAR immunotherapy. Recent work in our laboratory suggests that the quality of the immunological synapse (IS) can accurately predict CAR-modified cell efficacy (and toxicity) that can correlate with clinical outcomes. Here we review current efforts to develop a Synapse Predicts Efficacy (SPE) system for easy, rapid and cost-effective evaluation of CAR-modified immune cell immunotherapy. Ultimately, we hypothesize the conceptual basis and clinical application of SPE will serve as an important parameter in evaluating CAR immunotherapy and significantly advance precision cancer immunotherapy. [MediaObject not available: see fulltext.] Graphical abstract: Graphic abstract for manuscript CCAS-D-20-00136 by Liu, D., et al., 'The Role of Immunological Synapse in Predicting the Efficacy of Chimeric Antigen Receptor (CAR) Immunotherapy". The various branches of evaluating cancer immunotherapy metaphorically represented as a Rubik's cube. The development of a novel approach to predict the effectiveness of Chimeric Antigen Receptor (CAR)-modified cells by quantifying the quality of CAR IS will introduce a new parameter to the rapidly expanding field of cancer immunotherapy. Currently, no single parameter can predict the clinical outcome or efficacy of a specific type of CAR-modified cell. IS quality will serve as a quantifiable measure to evaluate CAR products and can be used in conjunction with other conventional parameters to form a composite clinical predictor. Much like a Rubik's cube has countless configurations, several methods and combinations of clinical metrics have arisen for evaluating the ability of a given immunotherapeutic strategy to treat cancer. The quality of IS depicting cancer immunotherapy is metaphorically expressed as a Rubik's cube. Each face/color represents one aspect of cancer therapy. Each grid in one face indicates one factor within that aspect of cancer therapy. For example, the green color represents the tumor microenvironment, and one out of the nine grids in the green color indicates suppressor cells (suppressors in green). Changes in one factor may completely alter the entire strategy of cancer therapy. However, the quality of IS (illuminated center red grid) makes the effectiveness of CAR immunotherapy predictable

Carbonic anhydrase enzymes regulate mast cell–mediated inflammation

Type 2 cytokine responses are necessary for the development of protective immunity to helminth parasites but also cause the inflammation associated with allergies and asthma. Recent studies have found that peripheral hematopoietic progenitor cells contribute to type 2 cytokine–mediated inflammation through their enhanced ability to develop into mast cells. In this study, we show that carbonic anhydrase (Car) enzymes are up-regulated in type 2–associated progenitor cells and demonstrate that Car enzyme inhibition is sufficient to prevent mouse mast cell responses and inflammation after Trichinella spiralis infection or the induction of food allergy–like disease. Further, we used CRISPR/Cas9 technology and illustrate that genetically editing Car1 is sufficient to selectively reduce mast cell development. Finally, we demonstrate that Car enzymes can be targeted to prevent human mast cell development. Collectively, these experiments identify a previously unrecognized role for Car enzymes in regulating mast cell lineage commitment and suggest that Car enzyme inhibitors may possess therapeutic potential that can be used to treat mast cell–mediated inflammation

Early innate immunity determines outcome of Mycobacterium tuberculosis pulmonary infection in rabbits

Background: Pulmonary infection of humans by Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), results in active disease in 5-10% of individuals, while asymptomatic latent Mtb infection (LTBI) is established in the remainder. The host immune responses that determine this differential outcome following Mtb infection are not fully understood. Using a rabbit model of pulmonary TB, we have shown that infection with the Mtb clinical isolate HN878 (a hyper-virulent W-Beijing lineage strain) leads to progressive cavitary disease similar to what is seen in humans with active TB. In contrast, infection with Mtb CDC1551 (a hyper-immunogenic clinical isolate) is efficiently controlled in rabbit lungs, with establishment of LTBI, which can be reactivated upon treatment with immune-suppressive drugs. We hypothesize that the initial interaction of Mtb with the cells of the host response in the lungs determine later outcome of infection. Results: To test this hypothesis, we used our rabbit model of pulmonary TB and infected the animals with Mtb HN878 or CDC1551. At 3 hours, with similar lung bacillary loads, HN878 infection caused greater accumulation of mononuclear and polymorphonuclear leukocytes (PMN) in the lungs, compared to animals infected with CDC1551. Using whole-genome microarray gene expression analysis, we delineated the early transcriptional changes in the lungs of HN878- or CDC1551-infected rabbits at this time and compared them to the differential response at 4 weeks of Mtb-infection. Our gene network and pathway analysis showed that the most significantly differentially expressed genes involved in the host response to HN878, compared to CDC1551, at 3 hours of infection, were components of the inflammatory response and STAT1 activation, recruitment and activation of macrophages, PMN, and fMLP (N-formyl-Methionyl-Leucyl-Phenylalanine)-stimulation. At 4 weeks, the CDC1551 bacillary load was significantly lower and the granulomatous response reduced compared to HN878 infection. Moreover, although inflammation was dampened in both Mtb infections at 4 weeks, the majority of the differentially expressed gene networks were similar to those seen at 3 hours. Conclusions: We propose that differential regulation of the inflammation-associated innate immune response and related gene expression changes seen at 3 hours determine the long term outcome of Mtb infection in rabbit lungs

Identification of infrastructure related risk factors, Deliverable 5.1 of the H2020 project SafetyCube

The present Deliverable (D5.1) describes the identification and evaluation of infrastructure related risk factors. It outlines the results of Task 5.1 of WP5 of SafetyCube, which aimed to identify and evaluate infrastructure related risk factors and related road safety problems by (i) presenting a taxonomy of infrastructure related risks, (ii) identifying “hot topics” of concern for relevant stakeholders and (iii) evaluating the relative importance for road safety outcomes (crash risk, crash frequency and severity etc.) within the scientific literature for each identified risk factor. To help achieve this, Task 5.1 has initially exploited current knowledge (e.g. existing studies) and, where possible, existing accident data (macroscopic and in-depth) in order to identify and rank risk factors related to the road infrastructure. This information will help further on in WP5 to identify countermeasures for addressing these risk factors and finally to undertake an assessment of the effects of these countermeasures. In order to develop a comprehensive taxonomy of road infrastructure-related risks, an overview of infrastructure safety across Europe was undertaken to identify the main types of road infrastructure-related risks, using key resources and publications such as the European Road Safety Observatory (ERSO), The Handbook of Road Safety Measures (Elvik et al., 2009), the iRAP toolkit and the SWOV factsheets, to name a few. The taxonomy developed contained 59 specific risk factors within 16 general risk factors, all within 10 infrastructure elements. In addition to this, stakeholder consultations in the form of a series of workshops were undertaken to prioritise risk factors (‘hot topics’) based on the feedback from the stakeholders on which risk factors they considered to be the most important or most relevant in terms of road infrastructure safety. The stakeholders who attended the workshops had a wide range of backgrounds (e.g. government, industry, research, relevant consumer organisations etc.) and a wide range of interests and knowledge. The identified ‘hot topics’ were ranked in terms of importance (i.e. which would have the greatest effect on road safety). SafetyCube analysis will put the greatest emphasis on these topics (e.g. pedestrian/cyclist safety, crossings, visibility, removing obstacles). To evaluate the scientific literature, a methodology was developed in Work Package 3 of the SafetyCube project. WP5 has applied this methodology to road infrastructure risk factors. This uniformed approach facilitated systematic searching of the scientific literature and consistent evaluation of the evidence for each risk factor. The method included a literature search strategy, a ‘coding template’ to record key data and metadata from individual studies, and guidelines for summarising the findings (Martensen et al, 2016b). The main databases used in the WP5 literature search were Scopus and TRID, with some risk factors utilising additional database searches (e.g. Google Scholar, Science Direct). Studies using crash data were considered highest priority. Where a high number of studies were found, further selection criteria were applied to ensure the best quality studies were included in the analysis (e.g. key meta-analyses, recent studies, country origin, importance). Once the most relevant studies were identified for a risk factor, each study was coded within a template developed in WP3. Information coded for each study included road system element, basic study information, road user group information, study design, measures of exposure, measures of outcomes and types of effects. The information in the coded templates will be included in the relational database developed to serve as the main source (‘back end’) of the Decision Support System (DSS) being developed for SafetyCube. Each risk factor was assigned a secondary coding partner who would carry out the control procedure and would discuss with the primary coding partner any coding issues they had found. Once all studies were coded for a risk factor, a synopsis was created, synthesising the coded studies and outlining the main findings in the form of meta-analyses (where possible) or another type of comprehensive synthesis (e.g. vote-count analysis). Each synopsis consists of three sections: a 2 page summary (including abstract, overview of effects and analysis methods); a scientific overview (short literature synthesis, overview of studies, analysis methods and analysis of the effects) and finally supporting documents (e.g. details of literature search and comparison of available studies in detail, if relevant). To enrich the background information in the synopses, in-depth accident investigation data from a number of sources across Europe (i.e. GIDAS, CARE/CADaS) was sourced. Not all risk factors could be enhanced with this data, but where it was possible, the aim was to provide further information on the type of crash scenarios typically found in collisions where specific infrastructure-related risk factors are present. If present, this data was included in the synopsis for the specific risk factor. After undertaking the literature search and coding of the studies, it was found that for some risk factors, not enough detailed studies could be found to allow a synopsis to be written. Therefore, the revised number of specific risk factors that did have a synopsis written was 37, within 7 infrastructure elements. Nevertheless, the coded studies on the remaining risk factors will be included in the database to be accessible by the interested DSS users. At the start of each synopsis, the risk factor is assigned a colour code, which indicates how important this risk factor is in terms of the amount of evidence demonstrating its impact on road safety in terms of increasing crash risk or severity. The code can either be Red (very clear increased risk), Yellow (probably risky), Grey (unclear results) or Green (probably not risky). In total, eight risk factors were given a Red code (e.g. traffic volume, traffic composition, road surface deficiencies, shoulder deficiencies, workzone length, low curve radius), twenty were given a Yellow code (e.g. secondary crashes, risks associated with road type, narrow lane or median, roadside deficiencies, type of junction, design and visibility at junctions) seven were given a Grey code (e.g. congestion, frost and snow, densely spaced junctions etc.). The specific risk factors given the red code were found to be distributed across a range of infrastructure elements, demonstrating that the greatest risk is spread across several aspects of infrastructure design and traffic control. However, four ‘hot topics’ were rated as being risky, which were ‘small work-zone length’, ‘low curve radius’, ‘absence of shoulder’ and ‘narrow shoulder’. Some limitations were identified. Firstly, because of the method used to attribute colour code, it is in theory possible for a risk factor with a Yellow colour code to have a greater overall magnitude of impact on road safety than a risk factor coded Red. This would occur if studies reported a large impact of a risk factor but without sufficient consistency to allocate a red colour code. Road safety benefits should be expected from implementing measures to mitigate Yellow as well as Red coded infrastructure risks. Secondly, findings may have been limited by both the implemented literature search strategy and the quality of the studies identified, but this was to ensure the studies included were of sufficiently high quality to inform understanding of the risk factor. Finally, due to difficulties of finding relevant studies, it was not possible to evaluate the effects on road safety of all topics listed in the taxonomy. The next task of WP5 is to begin identifying measures that will counter the identified risk factors. Priority will be placed on investigating measures aimed to mitigate the risk factors identified as Red. The priority of risk factors in the Yellow category will depend on why they were assigned to this category and whether or not they are a hot topic

Identification of road user related risk factors, deliverable 4.1 of the H2020 project SafetyCube.

Safety CaUsation, Benefits and Efficiency (SafetyCube) is a European Commission supported Horizon 2020 project with the objective of developing an innovative road safety Decision Support System (DSS). The DSS will enable policy-makers and stakeholders to select and implement the most appropriate strategies, measures, and cost-effective approaches to reduce casualties of all road user types and all severities. This document is the first deliverable (4.1) of work package 4 which is dedicated to identifying and assessing human related risk factors and corresponding countermeasures as well as their effect on road safety. The focus of deliverable 4.1 is on identification and assessment of risk factors and describes the corresponding operational procedure and corresponding outcomes. The following steps have been carried out: Identification of human related risk factors – creation of a taxonomy Consultation of relevant stakeholders and policy papers for identification of topic with high priority (‘hot topics’) Systematic literature search and selection of relevant studies on identified risk factors •Coding of studies •Analysis of risk factors on basis of coded studies •Synopses of risk factors, including accident scenarios The core output of this task are synopses of risk factors which will be available through the DSS. Within the synopses, each risk factor was analysed systematically on basis of scientific studies and is further assigned to one of four levels of risk (marked with a colour code). Essential information of the more than 180 included studies were coded and will also be available in the database of the DSS. Furthermore, the synopses contain theoretical background on the risk factor and are prepared in different sections with different levels of detail for an academic as well as a non-academic audience. These sections are readable independently. It is important to note that the relationship between road safety and road user related risk factors is a difficult task. For some risk factors the available studies focused more on conditions of the behaviour (in which situations the behaviour is shown or which groups are more likely to show this behaviour) rather than the risk factor itself. Therefore, it cannot be concluded that those risk factors that have not often been studied or have to rely more indirect and arguably weaker methodologies, e.g. self-reports , do not increase the chance of a crash occurring. The following analysed risk factors were assessed as ‘risky’, ‘probably risky’ or ‘unclear’. No risk factors were identified as ‘probably not risky’. Risky Probably risky Unclear • Influenced driving – alcohol • Influenced Driving – drugs (legal & illegal) • Speeding and inappropriate speed • Traffic rule violations – red light running • Distraction – cell phone use (hand held) • Distraction – cell phone use (hands free) • Distraction – cell phone use (texting) • Fatigue – sleep disorders – sleep apnea • Risk taking – overtaking • Risk taking – close following behaviour • Insufficient knowledge and skills • Functional impairment – cognitive impairment • Functional impairment – vision loss • Diseases and disorders – diabetes • Personal factors – sensation seeking • Personal factors – ADHD • Emotions – anger, aggression • Fatigue – Not enough sleep/driving while tired • Distraction – conversation with passengers • Distraction – outside of vehicle • Distraction – cognitive overload and inattention • Functional impairment – hearing loss (few studies) • Observation errors (few studies) • Distraction – music – entertainment systems (many studies, mixed results) • Distraction – operating devices (many studies, mixed results) The next step in SafetyCube’s WP4 is to identify and assess the effectiveness of measures and to establish a link to the identified risk factors. The work of this first task indicates a set of risk factors that should be centre of attention when identifying corresponding road safety measures (category ‘risky’)

Task-related enhancement in corticomotor excitability during haptic sensing with the contra- or ipsilateral hand in young and senior adults

<p>Abstract</p> <p>Background</p> <p>Haptic sensing with the fingers represents a unique class of manipulative actions, engaging motor, somatosensory and associative areas of the cortex while requiring only minimal forces and relatively simple movement patterns. Using transcranial magnetic stimulation (TMS), we investigated task-related changes in motor evoked potential (MEP) amplitude associated with unimanual haptic sensing in two related experiments. In Experiment I, we contrasted changes in the excitability of the hemisphere controlling the task hand in young and old adults under two trial conditions, i.e. when participants either touched a fine grating (<it>smooth trials</it>) or touched a coarse grating to detect its groove orientation (<it>grating trials</it>). In Experiment II, the same contrast between tasks was performed but with TMS applied over the hemisphere controlling the resting hand, while also addressing hemispheric (right vs. left) and age differences.</p> <p>Results</p> <p>In Experiment I, a main effect of <it>trial type </it>on MEP amplitude was detected (p = 0.001), MEPs in the task hand being ~50% larger during grating than smooth trials. No interaction with age was detected. Similar results were found for Experiment II, <it>trial type </it>having a large effect on MEP amplitude in the resting hand (p < 0.001) owing to selective increase in MEP size (~2.6 times greater) for grating trials. No interactions with age or side (right vs. left) were detected.</p> <p>Conclusions</p> <p>Collectively, these results indicate that adding a haptic component to a simple unilateral finger action can elicit robust corticomotor facilitation not only in the working hemisphere but also in the opposite hemisphere. The fact that this facilitation seems well preserved with age, when task difficulty is adjusted, has some potential clinical implications.</p

Inventory of assessed infrastructure risk factors and measures, Deliverable 5.4 of the H2020 project SafetyCube

Inventory of assessed infrastructure risk factors and measures, Deliverable 5.4 of the H2020 project SafetyCub

Profiling and Functional Analyses of MicroRNAs and Their Target Gene Products in Human Uterine Leiomyomas

Human uterine leiomyomas (ULM) are characterized by dysregulation of a large number of genes and non-coding regulatory microRNAs. In order to identify microRNA::mRNA associations relevant to ULM pathogenesis, we examined global correlation patterns between the altered microRNA expression and the predicted target genes in ULMs and matched myometria.Patterns of inverse association of microRNA with mRNA expression in ULMs revealed an involvement of multiple candidate pathways, including extensive transcriptional reprogramming, cell proliferation control, MAP kinase, TGF-beta, WNT, JAK/STAT signaling, remodeling of cell adhesion, and cell-cell and cell-matrix contacts. We further examined the correlation between the expression of the selected target gene protein products and microRNAs in thirty-six paired sets of leiomyomas and matched myometria. We found that a number of dysregulated microRNAs were inversely correlated with their targets at the protein level. The comparative genomic hybridization (CGH) in eight ULM patients revealed that partially shared deletions of two distinct chromosomal regions might be responsible for loss of cancer-associated microRNA expression and could thus contribute to the ULM pathogenesis via deregulation of target mRNAs. Last, we functionally tested the repressor effects of selected cancer-related microRNAs on their predicted target genes in vitro.We found that some but not all of the predicted and inversely correlated target genes in ULMs can be directly regulated by microRNAs in vitro. Our findings provide a broad overview of molecular events underlying the tumorigenesis of uterine ULMs and identify select genetic and regulatory events that alter microRNA expression and may play important roles in ULM pathobiology by positively regulating tumor growth while maintaining the non-invasive character of ULMs