miRNA-Mediated Fine Regulation of TLR-Induced M1 Polarization.

Macrophage polarization to the M1 spectrum is induced by bacterial cell wall components through stimulation of Toll-like family (TLR) receptors. By orchestrating the expression of relevant mediators of the TLR cascade, as well as associated pathways and feedback loops, macrophage polarization is coordinated to ensure an appropriate immune response. This is central to the successful control of pathogens and the maintenance of health. Macrophage polarization is known to be modulated at both the transcriptional and post-transcriptional levels. In recent years, the miRNA-based post-transcriptional regulation of M1 polarization has received increasing attention from the scientific community. Comparative studies have shown that TLR stimulation alters the miRNA profile of macrophages and that macrophages from the M1 or the M2 spectrum differ in terms of miRNAs expressed. Simultaneously, miRNAs are considered critical post-transcriptional regulators of macrophage polarization. In particular, miRNAs are thought to play a regulatory role in the switch between the early proinflammatory response and the resolution phase. In this review, we will discuss the current state of knowledge on the complex interaction of transcriptional and post-transcriptional regulatory mechanisms that ultimately determine the functionality of macrophages.

Ten-electron count rule for the binding of adsorbates on single-atom alloy catalysts.

Single-atom alloys have recently emerged as highly active and selective alloy catalysts. Unlike pure metals, single-atom alloys escape the well-established conceptual framework developed nearly three decades ago for predicting catalytic performance. Although this offers the opportunity to explore so far unattainable chemistries, this leaves us without a simple guide for the design of single-atom alloys able to catalyse targeted reactions. Here, based on thousands of density functional theory calculations, we reveal a 10-electron count rule for the binding of adsorbates on the dopant atoms, usually the active sites, of single-atom alloy surfaces. A simple molecular orbital approach rationalizes this rule and the nature of the adsorbate-dopant interaction. In addition, our intuitive model can accelerate the rational design of single-atom alloy catalysts. Indeed, we illustrate how the unique insights provided by the electron count rule help identify the most promising dopant for an industrially relevant hydrogenation reaction, thereby reducing the number of potential materials by more than one order of magnitude.

Transcriptom and miRNA data of PUFA-enriched stimulated murine macrophage and human endothelial cell lines.

Inflammation is associated with the adaptation of macrophages and endothelial cells, and the dysregulation of these differentiation processes has been directly linked to both acute and chronic disease states. As cells in constant contact with blood, macrophages and endothelial cells are also under the direct influence of immunomodulatory dietary components such as polyunsaturated fatty acids (PUFA). RNA sequencing analyses allow us to understand the global changes in gene expression occurring during cell differentiation, including both transcriptional (transcriptome) and post-transcriptional (miRNAs) levels. We generated a comprehensive RNA sequencing dataset of parallel transcriptome and miRNA profiles of PUFA-enriched and pro-inflammatory stimulated macrophages and endothelial cells aiming to uncover the underlying molecular mechanisms. PUFA concentrations and duration of supplementation were based on dietary ranges, allowing for metabolism and plasma membrane uptake of fatty acids. The dataset may serve as a resource to study transcriptional and post-transcriptional changes associated with macrophage polarisation and endothelial dysfunction in inflammatory settings and their modulation by omega-3 and omega-6 fatty acids.

Achieving Digital Catalysis: Strategies for Data Acquisition, Storage and Use.

Heterogeneous catalysis is an important area of research that generates data as intricate as the phenomenon itself. Complexity is inherently coupled to the function of the catalyst and advance in knowledge can only be achieved if this complexity is adequately captured and accounted for. This requires integration of experiment and theory, high data quality and quality control, close interdisciplinary collaboration, and sharing of data and metadata, which is facilitated by the application of joint data management strategies. This Viewpoint Article first discusses the potential of a digital transition in catalysis research. Then, a summary of the current status in terms of data infrastructure in heterogeneous catalysis is presented, defining the various types of (meta-) data, from catalyst synthesis to functional analysis. Finally, an already implemented working concept for local data acquisition and storage is introduced and the benefits and further development directions for catalysis data use and sharing are discussed.

Age and Comorbidity Burden of Patients Critically Ill with COVID-19 Affect Both Access to and Outcome of Ventilation Therapy in Intensive Care Units.

During the COVID-19 pandemic, large numbers of elderly, multimorbid people required treatment in intensive care units. This study investigated how the inherent patient factors age and comorbidity burden affected the treatment strategy and the outcome achieved. Retrospective analysis of data from intensive care patients enrolled in the Lean European Open Survey on SARS-CoV2-Infected Patients (LEOSS) cohort found that a patient's age and comorbidity burden in fact influenced their mortality rate and the use of ventilation therapy. Evidence showed that advanced age and multimorbidity were associated with the restrictive use of invasive ventilation therapies, particularly ECMO. Geriatric patients with a high comorbidity burden were clustered in the sub-cohort of non-ventilated ICU patients characterized by a high mortality rate. The risk of death generally increased with older age and accumulating comorbidity burden. Here, the more aggressive an applied procedure, the younger the age in which a majority of patients died. Clearly, geriatric, multimorbid COVID-19 patients benefit less from invasive ventilation therapies. This implies the need for a holistic approach to therapy decisions, taking into account the patient's wishes.

Invasiveness of Ventilation Therapy Is Associated to Prevalence of Secondary Bacterial and Fungal Infections in Critically Ill COVID-19 Patients.

Superinfections are a fundamental critical care problem, and their significance in severe COVID-19 cases needs to be determined. This study analyzed data from the Lean European Open Survey on SARS-CoV-2-Infected Patients (LEOSS) cohort focusing on intensive care patients. A retrospective analysis of patient data from 840 cases of COVID-19 with critical courses demonstrated that co-infections were frequently present and were primarily of nosocomial origin. Furthermore, our analysis showed that invasive therapy procedures accompanied an increased risk for healthcare-associated infections. Non-ventilated ICU patients were rarely affected by secondary infections. The risk of infection, however, increased even when non-invasive ventilation was used. A further, significant increase in infection rates was seen with the use of invasive ventilation and even more so with extracorporeal membrane oxygenation (ECMO) therapy. The marked differences among ICU techniques used for the treatment of COVID-19-induced respiratory failure in terms of secondary infection risk profile should be taken into account for the optimal management of critically ill COVID-19 patients, as well as for adequate antimicrobial therapy.

Molecular Mechanism of Cellular Membranes for Signal Transduction.

Membranes are central to cell function and crucial to life [...].

Observation and Characterization of Dicarbonyls on a RhCu Single-Atom Alloy.

Dicarbonyl species are ubiquitous on Rh/oxide catalysts and are known to form on Rh+ centers. However, dicarbonyl species have never been directly observed on single-atom alloys (SAAs) where the active site is metallic. Herein, using surface science and theoretical modeling, we provide evidence of dicarbonyl species at isolated Rh sites on a RhCu(100) SAA. This approach not only enables us to directly visualize dicarbonyl species at Rh sites but also demonstrates that the transition between the mono- and dicarbonyl configuration can be achieved by changing surface temperature and CO pressure. Density functional theory calculations further support the mono- and dicarbonyl assignments and provide evidence that these species should be stable on other SAA combinations. Together, these results provide a picture of the structure and energetics of both the mono- and dicarbonyl configurations on the RhCu(100) SAA surface and should aid with IR assignments on SAA nanoparticle catalysts.

miRNA-Mediated Priming of Macrophage M1 Differentiation Differs in Gram-Positive and Gram-Negative Settings.

A proper regulation of macrophage polarization is essential for the organism's health and pathogen control. Differentiation control is known to occur at the transcriptional as well as the posttranscriptional levels. The mechanisms involved, however, have not yet been fully elucidated. In this study, we co-cultured macrophages with viable Gram-positive and Gram-negative bacteria to mimic macrophage differentiation to the M1-like type in an inflammatory milieu. We found that Gram-positive stimulation resulted in increased expressions of miR-7a-5p, miR-148a-3p, miR-155-5p, and miR-351-5p. Of note, these miRNAs were found to target inhibitory mediators of the Rac1-PI3K-Akt pathway and the MyD88-dependent pathway. In contrast, Gram-negative stimulation-induced downregulation of miR-9-5p, miR-27b-3p, miR-93-5p, and miR-106b-5p is known to target key members of the Rac1-PI3K-Akt pathway and the MyD88-dependent pathway. These results, taken together, point to a fine-tuning of macrophage polarization by TLR-induced changes in macrophage miRNA profiles. Here, the miRNA-mediated priming of M1 differentiation seems to differ in the Gram-positive and Gram-negative settings in terms of the mechanism and miRNAs involved.

Loss of Endothelial Barrier Function in the Inflammatory Setting: Indication for a Cytokine-Mediated Post-Transcriptional Mechanism by Virtue of Upregulation of miRNAs miR-29a-3p, miR-29b-3p, and miR-155-5p.

Dysfunction of the endothelial barrier plays a central role in the pathogenesis of both acute and chronic inflammatory processes such as sepsis or atherosclerosis. Due to attenuation of endothelial cell contacts, there is an increased transfer of blood proteins and fluid into the surrounding tissue, which relates to edema formation and distribution disorders. However, the mechanisms underlying these responses are not fully understood. In this study, we used human endothelial cells to mimic the loss of barrier function in an inflammatory milieu. We found that a weakened endothelial barrier after cytokine stimulation was accompanied by a significantly changed transcriptome. Apparent was a depletion of mRNAs encoding cell adhesion molecules. Furthermore, we found that cytokine treatment of endothelial cells induced upregulation of miR-29a-3p, miR-29b-3p, and miR-155-5p. miRNAs are known to negatively affect stability and translational efficiency of target mRNAs. Remarkably, miR-29a-3p, miR-29b-3p, and miR-155-5p have already been described to target the mRNAs of central tight and adherent junction proteins including F11 receptor, claudin 1, ß-catenin, p120-catenin, and eplin. This taken together points to the existence of a posttranscriptional mechanism for expression inhibition of central adhesion proteins, which is triggered by inflammatory cytokines and mediated by miR-29a-3p, miR-29b-3p, and miR-155-5p.

Periodic Trends in Adsorption Energies around Single-Atom Alloy Active Sites.

Single-atom alloys (SAAs) make up a special class of alloy surface catalysts that offer well-defined, isolated active sites in a more inert metal host. The dopant sites are generally assumed to have little or no influence on the properties of the host metal, and transport of chemical reactants and products to and from the dopant sites is generally assumed to be facile. Here, by performing density functional theory calculations and surface science experiments, we identify a new physical effect on SAA surfaces, whereby adsorption is destabilized by ≤300 meV on host sites within the perimeter of the reactive dopant site. We identify periodic trends for this behavior and demonstrate a zone of exclusion around the reactive sites for a range of adsorbates and combinations of host and dopant metals. Experiments confirm an increased barrier for diffusion of CO toward the dopant on a RhCu SAA. This effect offers new possibilities for understanding and designing active sites with tunable energetic landscapes surrounding them.

The Reaction of CO2 with a Borylnitrene: Formation of an 3-Oxaziridinone.

The reaction of a borylnitrene with carbon dioxide is studied under cryogenic matrix isolation conditions. Photogenerated CatBN (Cat=catecholato) reacts with CO2 under formation of the cycloaddition product CatBNCO2 , a 3-oxaziridinone derivative, after photoexcitation (>550 nm). The product shows Fermi resonances between the CO stretching and ring deformation modes that cause unusual 13 C and 18 O isotopic shifts. A computational analysis of the 3-oxaziridinone shows this cyclic carbamate to be less strained than an α-lactone or an α-lactame.

Relative versus absolute RNA quantification: a comparative analysis based on the example of endothelial expression of vasoactive receptors.

BACKGROUND: In the present study, two distinct PCR methods were used for the quantification of genetic material and their results were compared: real-time-PCR (qPCR; relative quantification) and droplet digital PCR (ddPCR; absolute quantification). The comparison of the qPCR and the ddPCR was based on a stimulation approach of microvascular endothelial cells in which the effect of a pro-inflammatory milieu on the expression of vasoactive receptors was investigated. RESULTS: There was consistency in directions of effects for the majority of genes tested. With regard to the indicated dimension of the effects, the overall picture was more differentiated. It was striking that deviations were more pronounced if the measured values were on the extreme edges of the dynamic range of the test procedures. CONCLUSIONS: To obtain valid and reliable results, dilution series are recommended, which should be carried out initially. In case of ddPCR the number of copies per µl should be adjusted to the low three-digit range. With regard to qPCR it is essential that the stability and reliability of the reference genes used is guaranteed. Here, ddPCR offers the advantage that housekeeping genes are not required. Furthermore, an absolute quantification of the sample can be easily performed by means of ddPCR. Before using ddPCR, however, care should be taken to optimize the experimental conditions. Strict indications for this methodology should also be made with regard to economic and timing factors.

Inotropic agents and vasodilator strategies for the treatment of cardiogenic shock or low cardiac output syndrome.

BACKGROUND: Cardiogenic shock (CS) and low cardiac output syndrome (LCOS) are potentially life-threatening complications of acute myocardial infarction (AMI), heart failure (HF) or cardiac surgery. While there is solid evidence for the treatment of other cardiovascular diseases of acute onset, treatment strategies in haemodynamic instability due to CS and LCOS remains less robustly supported by the given scientific literature. Therefore, we have analysed the current body of evidence for the treatment of CS or LCOS with inotropic and/or vasodilating agents. This is the second update of a Cochrane review originally published in 2014. OBJECTIVES: Assessment of efficacy and safety of cardiac care with positive inotropic agents and vasodilator agents in CS or LCOS due to AMI, HF or after cardiac surgery. SEARCH METHODS: We conducted a search in CENTRAL, MEDLINE, Embase and CPCI-S Web of Science in October 2019. We also searched four registers of ongoing trials and scanned reference lists and contacted experts in the field to obtain further information. No language restrictions were applied. SELECTION CRITERIA: Randomised controlled trials (RCTs) enrolling patients with AMI, HF or cardiac surgery complicated by CS or LCOS. DATA COLLECTION AND ANALYSIS: We used standard methodological procedures according to Cochrane standards. MAIN RESULTS: We identified 19 eligible studies including 2385 individuals (mean or median age range 56 to 73 years) and three ongoing studies. We categorised studies into 11 comparisons, all against standard cardiac care and additional other drugs or placebo. These comparisons investigated the efficacy of levosimendan versus dobutamine, enoximone or placebo; enoximone versus dobutamine, piroximone or epinephrine-nitroglycerine; epinephrine versus norepinephrine or norepinephrine-dobutamine; dopexamine versus dopamine; milrinone versus dobutamine and dopamine-milrinone versus dopamine-dobutamine. All trials were published in peer-reviewed journals, and analyses were done by the intention-to-treat (ITT) principle. Eighteen of 19 trials were small with only a few included participants. An acknowledgement of funding by the pharmaceutical industry or missing conflict of interest statements occurred in nine of 19 trials. In general, confidence in the results of analysed studies was reduced due to relevant study limitations (risk of bias), imprecision or indirectness. Domains of concern, which showed a high risk in more than 50% of included studies, encompassed performance bias (blinding of participants and personnel) and bias affecting the quality of evidence on adverse events. All comparisons revealed uncertainty on the effect of inotropic/vasodilating drugs on all-cause mortality with a low to very low quality of evidence. In detail, the findings were: levosimendan versus dobutamine (short-term mortality: RR 0.60, 95% CI 0.36 to 1.03; participants = 1701; low-quality evidence; long-term mortality: RR 0.84, 95% CI 0.63 to 1.13; participants = 1591; low-quality evidence); levosimendan versus placebo (short-term mortality: no data available; long-term mortality: RR 0.55, 95% CI 0.16 to 1.90; participants = 55; very low-quality evidence); levosimendan versus enoximone (short-term mortality: RR 0.50, 0.22 to 1.14; participants = 32; very low-quality evidence; long-term mortality: no data available); epinephrine versus norepinephrine-dobutamine (short-term mortality: RR 1.25; 95% CI 0.41 to 3.77; participants = 30; very low-quality evidence; long-term mortality: no data available); dopexamine versus dopamine (short-term mortality: no deaths in either intervention arm; participants = 70; very low-quality evidence; long-term mortality: no data available); enoximone versus dobutamine (short-term mortality RR 0.21; 95% CI 0.01 to 4.11; participants = 27; very low-quality evidence; long-term mortality: no data available); epinephrine versus norepinephrine (short-term mortality: RR 1.81, 0.89 to 3.68; participants = 57; very low-quality evidence; long-term mortality: no data available); and dopamine-milrinone versus dopamine-dobutamine (short-term mortality: RR 1.0, 95% CI 0.34 to 2.93; participants = 20; very low-quality evidence; long-term mortality: no data available). No information regarding all-cause mortality were available for the comparisons milrinone versus dobutamine, enoximone versus piroximone and enoximone versus epinephrine-nitroglycerine. AUTHORS' CONCLUSIONS: At present, there are no convincing data supporting any specific inotropic or vasodilating therapy to reduce mortality in haemodynamically unstable patients with CS or LCOS. Considering the limited evidence derived from the present data due to a high risk of bias and imprecision, it should be emphasised that there is an unmet need for large-scale, well-designed randomised trials on this topic to close the gap between daily practice in critical care of cardiovascular patients and the available evidence. In light of the uncertainties in the field, partially due to the underlying methodological flaws in existing studies, future RCTs should be carefully designed to potentially overcome given limitations and ultimately define the role of inotropic agents and vasodilator strategies in CS and LCOS.

Polyunsaturated Fatty Acids of Both the Omega-3 and the Omega-6 Family Abrogate the Cytokine-Induced Upregulation of miR-29a-3p by Endothelial Cells.

Cellular processes fundamentally depend on protein expression control. At this, protein expression is regulated on the transcriptional and the post-transcriptional level. PUFAs are already known to affect gene transcription. The present study was conducted to answer the question whether PUFAs are also able to impact on the miRNA-mediated post-transcriptional fine-tuning of mRNA copy numbers. To this end, cellular miRNA profiles were screened by means of next-generation sequencing and NanoString analysis to compare PUFA-enriched to unsupplemented endothelial cells exposed to an inflammatory milieu. Validation took place by droplet digital PCR, allowing for an absolute quantification of RNA copy numbers. The analyses revealed that the stimulation-induced upregulation of miR-29a-3p is blocked by PUFA enrichment of endothelial cells. What is more, mRNA copy numbers of miR-29a-3p targets, namely the coagulation factors PAI-1, TF, and vWF, as well as the proinflammatory cytokines IL-1ß, IL-6, and IL-8, were reduced in PUFA-enriched endothelial cells compared to unsupplemented cells, counteracting the stimulatory effect of an inflammatory environment. These data hint toward a new mechanism of action by which PUFAs modulate the functionality of endothelial cells. Apparently, the inflammation-modulating properties of PUFAs are also mediated at the post-transcriptional level.

Unusual Nitrene Oxidation Product Formation by Metathesis Involving the Dioxygen O-O and Borylnitrene B-N Bonds.

The reaction of dioxygen with nitrenes can have significant energy barriers, although both reactants are triplet diradicals and the formation of nitroso-O-oxides is spin-allowed. By means of matrix-isolation infrared spectroscopy in solid argon, nitrogen, and neon, and through high-level computational quantum chemistry, it is shown herein that a 3-nitreno-1,3,2-benzodioxaborole CatBN (Cat=catecholato) reacts with dioxygen under cryogenic conditions thermally at temperatures as low as 7 K to produce two distinct products, an anti-nitroso-O-oxide and a nitritoborane CatBONO. The computed barriers for the formation of nitroso-O-oxide isomers are very low. Whereas anti-nitroso-O-oxide is kinetically trapped, its bisected isomer has a very low barrier for metathesis, yielding the CatBO+NO radicals in a strongly exothermic reaction; these radicals can combine under matrix-isolation conditions to give nitritoborane CatBONO. The trapped isomer, anti-nitroso-O-oxide, can form the nitritoborane CatBONO only after photoexcitation, possibly involving isomerization to the bisected isomer of anti-nitroso-O-oxide.

Sepsis Diagnostics: Intensive Care Scoring Systems Superior to MicroRNA Biomarker Testing.

Sepsis represents a serious medical problem accounting for numerous deaths of critically ill patients in intensive care units (ICUs). An early, sensitive, and specific diagnosis is considered a key element for improving the outcome of sepsis patients. In addition to classical laboratory markers, ICU scoring systems and serum miRNAs are discussed as potential sepsis biomarkers. In the present prospective observational study, the suitability of miRNAs in sepsis diagnosis was tested based on proper validated and normalized data (i.e., absolute quantification by means of Droplet Digital PCR (ddPCR)) in direct comparison to classical sepsis markers and ICU scores within the same patient cohort. Therefore, blood samples of septic intensive care patients (n = 12) taken at day of admission at ICU were compared to non-septic intensive care patients (n = 12) and a healthy control group (n = 12). Our analysis indicates that all tested biomarkers have only a moderate informative power and do not allow an unequivocal differentiation between septic and non-septic ICU patients. In conclusion, there is no standalone laboratory parameter that enables a reliable diagnosis of sepsis. miRNAs are not superior to classical parameters in this respect. It seems recommendable to measure multiple parameters and scores and to interpret them with regard to the clinical presentation.

Author Correction: Identification of stably expressed housekeeping miRNAs in endothelial cells and macrophages in an inflammatory setting.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Identification of stably expressed housekeeping miRNAs in endothelial cells and macrophages in an inflammatory setting.

Reliable quantification of miRNA expression by qRT-PCR crucially depends on validated housekeepers for data normalization. Here we present thoroughly tested miRNAs eligible as references in immunological studies utilizing endothelial cells and macrophages, respectively. Endothelial cells (cell line: TIME) and macrophages (cell line: RAW264.7) were treated with various pro- and anti-inflammatory mediators (cytokines, LPS, unsaturated fatty acids) given as either single substances or in combination. Isolated RNA was screened for stably expressed miRNAs by next generation sequencing. Housekeeper candidates were thereafter validated by means of two independent quantification techniques: qRT-PCR for relative quantification and ddPCR for absolute quantification. Both methods consistently confirmed the suitability of let-7g-5p, let-7i-5p, miR-127-3p and miR-151a-5p in cytokine/fatty acid-treated TIME and miR-16-5p, miR-27b-3p, miR-103a-3p and miR-423-3p in LPS/fatty acid-treated RAW264.7, respectively as housekeeping miRNAs. With respect to abundancy and over all expression stability the miRNAs miR-151a-5p (cell line: TIME) as well as miR-27b-3p and miR-103a-3p (cell line: RAW264.7) can be particularly recommended for normalization of qRT-PCR data.