The prospective isolation of purified stem cell populations has dramatically altered the field of stem cell biology, and it has been a major focus of research across tissues in different organisms. ...Muscle stem cells (MuSCs) are now among the most intensely studied stem cell populations in mammalian systems, and the prospective isolation of these cells has allowed cellular and molecular characterizations that were not dreamed of a decade ago. In this protocol, we describe how to isolate MuSCs from limb muscles of adult mice by fluorescence-activated cell sorting (FACS). We provide a detailed description of the physical and enzymatic dissociation of mononucleated cells from limb muscles, a procedure that is essential in order to maximize cell yield. We also describe a FACS-based method that is used subsequently to obtain highly pure populations of either quiescent or activated MuSCs (VCAM(+)CD31(-)CD45(-)Sca1(-)). The isolation process takes ∼5-6 h to complete. The protocol also allows for the isolation of endothelial cells, hematopoietic cells and mesenchymal stem cells from muscle tissue.
Background
Current guidelines for the treatment of adult severe traumatic brain injury (sTBI) consist of high-quality evidence reports, but they are no longer accompanied by management protocols, as ...these require expert opinion to bridge the gap between published evidence and patient care. We aimed to establish a modern sTBI protocol for adult patients with both intracranial pressure (ICP) and brain oxygen monitors in place.
Methods
Our consensus working group consisted of 42 experienced and actively practicing sTBI opinion leaders from six continents. Having previously established a protocol for the treatment of patients with ICP monitoring alone, we addressed patients who have a brain oxygen monitor in addition to an ICP monitor. The management protocols were developed through a Delphi-method-based consensus approach and were finalized at an in-person meeting.
Results
We established three distinct treatment protocols, each with three tiers whereby higher tiers involve therapies with higher risk. One protocol addresses the management of ICP elevation when brain oxygenation is normal. A second addresses management of brain hypoxia with normal ICP. The third protocol addresses the situation when both intracranial hypertension and brain hypoxia are present. The panel considered issues pertaining to blood transfusion and ventilator management when designing the different algorithms.
Conclusions
These protocols are intended to assist clinicians in the management of patients with both ICP and brain oxygen monitors but they do not reflect either a standard-of-care or a substitute for thoughtful individualized management. These protocols should be used in conjunction with recommendations for basic care, management of critical neuroworsening and weaning treatment recently published in conjunction with the Seattle International Brain Injury Consensus Conference.
Machine learning may be useful to characterize cardiovascular risk, predict outcomes, and identify biomarkers in population studies.
To test the ability of random survival forests, a machine learning ...technique, to predict 6 cardiovascular outcomes in comparison to standard cardiovascular risk scores.
We included participants from the MESA (Multi-Ethnic Study of Atherosclerosis). Baseline measurements were used to predict cardiovascular outcomes over 12 years of follow-up. MESA was designed to study progression of subclinical disease to cardiovascular events where participants were initially free of cardiovascular disease. All 6814 participants from MESA, aged 45 to 84 years, from 4 ethnicities, and 6 centers across the United States were included. Seven-hundred thirty-five variables from imaging and noninvasive tests, questionnaires, and biomarker panels were obtained. We used the random survival forests technique to identify the top-20 predictors of each outcome. Imaging, electrocardiography, and serum biomarkers featured heavily on the top-20 lists as opposed to traditional cardiovascular risk factors. Age was the most important predictor for all-cause mortality. Fasting glucose levels and carotid ultrasonography measures were important predictors of stroke. Coronary Artery Calcium score was the most important predictor of coronary heart disease and all atherosclerotic cardiovascular disease combined outcomes. Left ventricular structure and function and cardiac troponin-T were among the top predictors for incident heart failure. Creatinine, age, and ankle-brachial index were among the top predictors of atrial fibrillation. TNF-α (tissue necrosis factor-α) and IL (interleukin)-2 soluble receptors and NT-proBNP (N-Terminal Pro-B-Type Natriuretic Peptide) levels were important across all outcomes. The random survival forests technique performed better than established risk scores with increased prediction accuracy (decreased Brier score by 10%-25%).
Machine learning in conjunction with deep phenotyping improves prediction accuracy in cardiovascular event prediction in an initially asymptomatic population. These methods may lead to greater insights on subclinical disease markers without apriori assumptions of causality.
URL: http://www.clinicaltrials.gov. Unique identifier: NCT00005487.
A flurry of recently published studies indicates that both insects and birds have experienced wide-scale population declines in the last several decades. Curiously, whether insect and bird declines ...are causally linked has received little empirical attention. Here, we hypothesize that insect declines are an important factor contributing to the decline of insectivorous birds. We further suggest that insect populations essential to insectivorous birds decline whenever nonnative lumber, ornamental, or invasive plant species replace native plant communities. We support our hypothesis by reviewing studies that show (1) due to host plant specialization, insect herbivores typically do poorly on non-native plants; (2) birds are often food limited; (3) populations of insectivorous bird species fluctuate with the supply of essential insect prey; (4) not all arthropod prey support bird reproduction equally well; and (5) terrestrial birds for which insects are an essential source of food have declined by 2.9 billion individuals over the last 50 years, while terrestrial birds that do not depend on insects during their life history have gained by 26.2 million individuals, a 111-fold difference. Understanding the consequences of insect declines, particularly as they affect charismatic animals like birds, may motivate land managers, homeowners, and restoration ecologists to take actions that reverse these declines by favoring the native plant species that support insect herbivores most productively. LAY SUMMARY There may be a link between declining insect populations and bird population declines. Bird populations are often limited by the amount of insect food available to them. Most of the bird species that have declined in the last 50 years are those that depend on insects for food. One cause of insect declines is the widespread use of non-native plants in forestry and horticulture that do not support insects. The loss of insects, birds, and other forms of life is important because those are the species that run the ecosystems that support humans. One solution to this problem is to increase the populations of the insects that birds depend on by increasing our use of native plants in managed landscapes.
The immune microenvironment of hepatocellular carcinoma (HCC) is poorly characterized. Combining two single-cell RNA sequencing technologies, we produced transcriptomes of CD45+ immune cells for HCC ...patients from five immune-relevant sites: tumor, adjacent liver, hepatic lymph node (LN), blood, and ascites. A cluster of LAMP3+ dendritic cells (DCs) appeared to be the mature form of conventional DCs and possessed the potential to migrate from tumors to LNs. LAMP3+ DCs also expressed diverse immune-relevant ligands and exhibited potential to regulate multiple subtypes of lymphocytes. Of the macrophages in tumors that exhibited distinct transcriptional states, tumor-associated macrophages (TAMs) were associated with poor prognosis, and we established the inflammatory role of SLC40A1 and GPNMB in these cells. Further, myeloid and lymphoid cells in ascites were predominantly linked to tumor and blood origins, respectively. The dynamic properties of diverse CD45+ cell types revealed by this study add new dimensions to the immune landscape of HCC.
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•Two scRNA-seq platforms reveal a high-resolution dynamic immune landscape in HCC•LAMP3+ DCs, arising from cDCs, can migrate from tumors to hepatic lymph nodes•Paired ligand-receptor analyses implicate the regulation of lymphocytes by LAMP3+ DCs•Macrophage subsets in tumors show distinct states and potentials to egress to ascites
Integrated single-cell RNA sequencing technologies and bioinformatics approaches reveal a high-resolution immune landscape of hepatocellular carcinoma, identifying inflammatory signatures and functional states of myeloid cells as well as predictions of complex cell-cell interactions.
Retinal ganglion cells (RGCs) are frequently divided into functional types by their ability to extract and relay specific features from a visual scene, such as the capacity to discern local or global ...motion, direction of motion, stimulus orientation, contrast or uniformity, or the presence of large or small objects. Here we introduce three previously uncharacterized, nondirection-selective ON-OFF RGC types that represent a distinct set of feature detectors in the mouse retina. The three high-definition (HD) RGCs possess small receptive-field centers and strong surround suppression. They respond selectively to objects of specific sizes, speeds, and types of motion. We present comprehensive morphological characterization of the HD RGCs and physiological recordings of their light responses, receptive-field size and structure, and synaptic mechanisms of surround suppression. We also explore the similarities and differences between the HD RGCs and a well characterized RGC with a comparably small receptive field, the local edge detector, in response to moving objects and textures. We model populations of each RGC type to study how they differ in their performance tracking a moving object. These results, besides introducing three new RGC types that together constitute a substantial fraction of mouse RGCs, provide insights into the role of different circuits in shaping RGC receptive fields and establish a foundation for continued study of the mechanisms of surround suppression and the neural basis of motion detection.
The output cells of the retina, retinal ganglion cells (RGCs), are a diverse group of ∼40 distinct neuron types that are often assigned "feature detection" profiles based on the specific aspects of the visual scene to which they respond. Here we describe, for the first time, morphological and physiological characterization of three new RGC types in the mouse retina, substantially augmenting our understanding of feature selectivity. Experiments and modeling show that while these three "high-definition" RGCs share certain receptive-field properties, they also have distinct tuning to the size, speed, and type of motion on the retina, enabling them to occupy different niches in stimulus space.
Organophosphonate-based nerve agents, such as VX, Sarin (GB), and Soman (GD), are among the most toxic chemicals to humankind. Recently, we have shown that Zr-based metal–organic frameworks (Zr-MOFs) ...can effectively catalyze the hydrolysis of these toxic chemicals for diminishing their toxicity. On the other hand, utilizing these materials in powder form is not practical, and developing scalable and economical processes for integrating these materials onto fibers is crucial for protective gear. Herein, we report a scalable, template-free, and aqueous solution-based synthesis strategy for the production of Zr-MOF-coated textiles. Among all MOF/fiber composites reported to date, the MOF-808/polyester fibers exhibit the highest rates of nerve agent hydrolysis. Moreover, such highly porous fiber composites display significantly higher protection time compared to that of its parent fabric for a mustard gas simulant, 2-chloroethyl ethyl sulfide (CEES). A decreased diffusion rate of toxic chemicals through the MOF layer can provide time needed for the destruction of the harmful species.
Allylsilanes can be regioselectively transformed into the corresponding 3-silylfluorohydrin in good yield using a sequence of epoxidation followed by treatment with HF·Et3N with or without isolation ...of the intermediate epoxide. Various silicon-substitutions are tolerated, resulting in a range of 2-fluoro-3-silylpropan-1-ol products from this method. Whereas other fluorohydrin syntheses by epoxide opening using HF·Et3N generally require more forcing conditions (e.g., higher reaction temperature), opening of allylsilane-derived epoxides with this reagent occurs at room temperature. We attribute this rate acceleration along with the observed regioselectivity to a β-silyl effect that stabilizes a proposed cationic intermediate. The use of enantioenriched epoxides indicates that both SN1- and SN2-type mechanisms may be operable depending on substitution at silicon. Conformational analysis by NMR and theory along with a crystal structure obtained by X-ray diffraction points to a preference for silicon and fluorine to be proximal to one another in the products, perhaps favored due to electrostatic interactions.
Rapid room-temperature synthesis of metal–organic frameworks (MOFs) is highly desired for industrial implementation and commercialization. Here we find that a (Zn,Cu) hydroxy double salt (HDS) ...intermediate formed in situ from ZnO particles or thin films enables rapid growth (<1 min) of HKUST-1 (Cu3(BTC)2) at room temperature. The space-time-yield reaches >3 × 104 kg·m–3·d–1, at least 1 order of magnitude greater than any prior report. The high anion exchange rate of (Zn,Cu) hydroxy nitrate HDS drives the ultrafast MOF formation. Similarly, we obtained Cu-BDC, ZIF-8, and IRMOF-3 structures from HDSs, demonstrating synthetic generality. Using ZnO thin films deposited via atomic layer deposition, MOF patterns are obtained on pre-patterned surfaces, and dense HKUST-1 coatings are grown onto various form factors, including polymer spheres, silicon wafers, and fibers. Breakthrough tests show that the MOF-functionalized fibers have high adsorption capacity for toxic gases. This rapid synthesis route is also promising for new MOF-based composite materials and applications.
Abstract
The scope and purpose of this work is 2-fold: to synthesize the available evidence and to translate it into recommendations. This document provides recommendations only when there is ...evidence to support them. As such, they do not constitute a complete protocol for clinical use. Our intention is that these recommendations be used by others to develop treatment protocols, which necessarily need to incorporate consensus and clinical judgment in areas where current evidence is lacking or insufficient. We think it is important to have evidence-based recommendations to clarify what aspects of practice currently can and cannot be supported by evidence, to encourage use of evidence-based treatments that exist, and to encourage creativity in treatment and research in areas where evidence does not exist. The communities of neurosurgery and neuro-intensive care have been early pioneers and supporters of evidence-based medicine and plan to continue in this endeavor. The complete guideline document, which summarizes and evaluates the literature for each topic, and supplemental appendices (A-I) are available online at https://www.braintrauma.org/coma/guidelines.
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