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Leonie M Becker,1,2 Aart Jan W Teunissen,2 Joseph SHA Koopman2 1Department of Cardiology, St. Antonius Ziekenhuis, Nieuwegein, The Netherlands; 2Department of Anaesthesiology, Maasstad Ziekenhuis, Rotterdam, The NetherlandsCorrespondence: Leonie M Becker, Department of Cardiology, St. Antonius Hospital, Koekoekslaan 1, Nieuwegein, 3435 CM, The Netherlands, Tel +31 88 320 09 22, Email l.becker@antoniusziekenhuis.nlAbstract: The addition of morphine to neuraxial anaesthesia leads to improved postoperative analgesia and lower opioid consumption, but is often accompanied by pruritus. Studies on preventing or treating pruritus show contradictory results. Our objective was to identify effective drugs for the prevention or treatment of pruritus by a scoping review of clinical trials. A systematic literature search was conducted in PubMed, Embase and Web of Science. We identified clinical trials investigating the prevention or treatment of neuraxial morphine-induced pruritus in adults. Systematic reviews and meta-analyses were screened for eligible studies. One-hundred-and-four articles were included covering 13 pharmacological groups. We conclude that dopamine antagonists, μ-opioid agonist/antagonists and neuraxial or orally administered μ-opioid antagonists prevent pruritus caused by neuraxial morphine regardless of the timing of administration. In the reviewed literature, 5HT3-antagonists prevent neuraxial morphine-induced pruritus when administered before morphine administration. For the treatment of neuraxial morphine-induced pruritus, only nalbuphine appears to be consistently effective. More research is needed to find the most effective doses and the optimal timing of the effective medication.Keywords: pruritus, prevention, treatment, morphine, epidural, spinal

Joyce Peper,1,2,* Leonie M Becker,1,2,* Jan-Peter van Kuijk,1 Tim Leiner,2 Martin J Swaans1 1Department of Cardiology, St. Antonius Hospital, Nieuwegein, The Netherlands; 2Department of Radiology, University Medical Center Utrecht, Utrecht, The Netherlands*These authors contributed equally to this workCorrespondence: Joyce PeperDepartment of Cardiology, St. Antonius Hospital, Koekoekslaan 1, Nieuwegein, 3435 CM, The NetherlandsTel +31 88 320 09 22Email j.peper@antoniusziekenhuis.nlAbstract: The aim of this review was to discuss the current practice and patient selection for invasive FFR, new techniques to estimate invasive FFR and future of coronary physiology tests. We elaborate on the indication and application of FFR and on the contraindications and concerns in certain patient populations.Keywords: coronary artery disease, fractional flow reserve, patient selection, revascularisation

Objectives: Screening for obstructive coronary artery disease (CAD) with coronary computed tomography angiography (CCTA) could prevent unnecessary invasive coronary angiography (ICA) procedures during work-up for trans-catheter aortic valve implantation (TAVI). CT-derived fractional flow reserve (CT-FFR) improves CCTA accuracy in chest pain patients. However, its reliability in the TAVI population is unknown. This systematic review and meta-analysis assesses CCTA and CT-FFR in TAVI candidates., Methods: PubMed, Embase and Web of Science were searched for studies regarding CCTA and/or CT-FFR in TAVI candidates. Primary endpoint was correct identification and rule-out of obstructive CAD. Results were pooled in a meta-analysis., Results: Thirty-four articles were part of the meta-analysis, reporting results for CCTA and CT-FFR in 7235 and 1269 patients, respectively. Reference standard was mostly anatomical severity of CAD. At patient level, pooled CCTA sensitivity was 94.0% and specificity 72.4%. CT-FFR sensitivity was 93.2% and specificity 70.3% with substantial variation between studies. However, in studies that compared both, CT-FFR performed better than CCTA. Sensitivity of CCTA versus CT-FFR was 74.9% versus 83.9%, and specificity was 65.5% versus 89.8%., Conclusions: Negative CCTA accurately rules out CAD in the TAVI population. CCTA could lead to significant reduction in pre-TAVI ICA, but false positives remain high. Diagnostic accuracy of CT-FFR was comparable to that of CCTA in our meta-analyses, but in studies performing a direct comparison, CT-FFR performed better than CCTA. However, as most studies were small and used CT-FFR software exclusively available for research, a large study on CT-FFR in TAVI work-up using commercially available CT-FFR software would be appropriate before considering routine implementation., Key Points: Question Coronary artery disease (CAD) screening with invasive coronary angiography before trans-catheter aortic valve implantation (TAVI) is often retrospectively unnecessary, revealing no obstructive CAD. Findings Coronary CTA ruled out CAD in approximately half of TAVI candidates. CT-derived fractional flow reserve (CT-FFR) performed similarly overall but better than coronary CTA in direct comparison. Clinical relevance Addition of coronary CTA to TAVI planning-CT to screen for obstructive CAD could reduce negative invasive coronary angiographies in TAVI work-up. CT-FFR could reduce false-positive coronary CTA results, improving its gatekeeper function in this population, but more data is necessary., Competing Interests: Compliance with ethical standards. Guarantor: The scientific guarantor of this publication is Dr M.J. Swaans. Conflict of interest: The authors of this manuscript declare no relationships with any companies, whose products or services may be related to the subject matter of the article. Statistics and biometry: One of the authors has significant statistical expertise (Joyce Peper). Informed consent: Written informed consent was not required for this study because this is a systematic review and meta-analysis on existing literature. Ethical approval: Institutional Review Board approval was not required because this is a systematic review and meta-analysis on existing literature. Study subjects or cohorts overlap: As this is a systematic review and meta-analysis on existing literature, all included subjects have been previously reported in published articles. Methodology: Meta-analysis, (© 2024. The Author(s), under exclusive licence to European Society of Radiology.)

Competing Interests: MJS declares a ZonMW Efficacy Studies grant (2021) for the iCORONARY trial (Improving the Cost-effectiveness of Coronary Artery Disease Diagnosis) and a ZonMW Promising Care grant (2022) for the TRACE-NL trial (Evaluation of the safety, efficacy and cost-effectiveness of transcatheter tricuspid valve repair in patients with severe tricuspid regurgitation in the Netherlands); and has received consultancy fees from Abbott Vascular, Bioventrix, Boston Scientific, Cardiac Dimensions, Edwards Lifesciences, GE Healthcare, Medtronic, Philips Healthcare, P&F, and Siemens Healthcare. LMB declares no competing interests.

Computed tomography-derived fractional flow reserve (CT-FFR) enhances the specificity of coronary computed tomography angiography (CCTA) to that of the most specific non-invasive imaging techniques, while maintaining high sensitivity in stable coronary artery disease (CAD). As gatekeeper for invasive coronary angiography (ICA), use of CT-FFR results in a significant reduction of negative ICA procedures and associated costs and complications, without increasing cardiovascular events. It is expected that CT-FFR algorithms will continue to improve, regarding accuracy and generalisability, and that introduction of new features will allow further treatment guidance and reduced invasive diagnostic testing. Advancements in CCTA quality and artificial intelligence (AI) are starting to unfold the incremental diagnostic and prognostic capabilities of CCTA's attenuation-based images in CAD, with future perspectives promising additional CCTA parameters which will enable non-invasive assessment of myocardial ischaemia as well as CAD activity and future cardiovascular risk. This review discusses practical application, interpretation and impact of CT-FFR on patient care, and how this ties into the CCTA 'one stop shop' for coronary assessment and patient prognosis. In this light, selective adoption of the most promising, objective and reproducible techniques and algorithms will yield maximal diagnostic value of CCTA without overcomplicating patient management and guideline recommendations., (© 2024. The Author(s).)

Growing evidence highlights the importance of tumor endothelial cells (TECs) in the tumor microenvironment (TME) for promoting tumor growth and evading immune responses. Immunomodulatory endothelial cells (IMECs) represent a distinct plastic phenotype of ECs that exerts the ability to modulate immunity in health and disease. This review discusses our current understanding of IMECs in cancer biology, scrutinizing insights from single-cell reports to compare their characteristics and function dynamics across diverse tumor types, conditions, and species. We investigate possible implications of exploiting IMECs in the context of cancer treatment, particularly examining their influence on the efficacy of existing therapies and the potential to leverage them as targets in optimizing immunotherapeutic strategies., Competing Interests: Declaration of interests None are declared by the authors., (Copyright © 2024. Published by Elsevier Inc.)

Carcinomas are associated with metastasis to specific organs while sparing others. Breast cancer presents with lung metastasis but rarely kidney metastasis. Using this difference as an example, we queried the mechanism(s) behind the proclivity for organ-specific metastasis. We used spontaneous and implant models of metastatic mammary carcinoma coupled with inflammatory tissue fibrosis, single-cell sequencing analyses and functional studies to unravel the causal determinants of organ-specific metastasis. Here we show that lung metastasis is facilitated by angiopoietin 2 (Ang2)-mediated suppression of lung-specific endothelial tight junction protein Claudin 5, which is augmented by the inflammatory fibrotic microenvironment and prevented by anti-Ang2 blocking antibodies, while kidney metastasis is prevented by non-Ang2-responsive Claudins 2 and 10. Suppression of Claudins 2 and 10 was sufficient to induce the emergence of kidney metastasis. This study illustrates the influence of organ-specific vascular heterogeneity in determining organotropic metastasis, independent of cancer cell-intrinsic mechanisms., (© 2024. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Purpose: The anterior stability of reverse total shoulder arthroplasty is affected by multiple factors. However, the effect of glenosphere inclination on stability has rarely been investigated, which is what this study aims to look into., Methods: Reverse shoulder arthroplasty was performed on 15 cadaveric human shoulders. The anterior dislocation forces and range of motion in internal rotation in the glenohumeral joint (primary measured parameters) were tested in a shoulder simulator in different arm positions and implant configurations, as well as with a custom-made 10° inferiorly inclined glenosphere. The inclination and retroversion of the baseplate as well as the distance between the glenoid and coracoid tip in two planes (secondary measured parameters) were evaluated on CT scans., Results: In biomechanical testing, the custom-made inclined glenosphere showed no significant influence on anterior stability other than glenoid lateralisation over all arm positions as well as the neck-shaft angle in two arm positions. The 6 mm lateralised glenosphere reduced internal rotation at 30° and 60° of glenohumeral abduction. In 30° of glenohumeral abduction, joint stability was increased using the 155° epiphysis compared with the 145° epiphysis. The mean inclination was 16.1°. The inclination was positively, and the distance between the glenoid and coracoid tip in the anterior-to-posterior direction was negatively correlated with anterior dislocation forces., Conclusions: The custom-made inferiorly inclined glenosphere did not influence anterior stability, but baseplate inclination itself had a significant effect on stability., (© 2024. The Author(s).)

Angiogenesis and lymphangiogenesis, the formation of new blood or lymphatic vessels, respectively, from preexisting vasculature is essential during embryonic development, but also occurs during tissue repair and in pathological conditions (cancer; ocular disease; ischemic, infectious and inflammatory disorders), which are all characterized to a certain extent by inflammatory conditions. Hence, a rapid, inexpensive, feasible / technically easy, reliable assay of inflammation-induced (lymph-)angiogenesis is highly valuable. In this context, the corneal thermal cauterization assay in mice is a simple, low-cost, reproducible, insightful and labor-saving assay to gauge the role of inflammation in angiogenesis and lymphangiogenesis. However, to the best of our knowledge, there is no standardized protocol to perform this assay. Here, we provide a step-by-step description of the model's procedures, which include:•The thermal cauterization of the corneas,•Enucleation and dissection of the corneas,•Subsequent immunofluorescence staining of the neovasculature, and morphometric analysis. We also discuss ethical considerations and aspects related to animal welfare guidelines. Altogether, this paper will help to increase the reproducibility of the corneal thermal cauterization model and facilitate its use for angiogenesis and lymphangiogenesis research., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors. Published by Elsevier B.V.)

Magic-angle spinning (MAS) nuclear magnetic resonance (NMR) is establishing itself as a powerful method for the characterization of protein dynamics at the atomic scale. We discuss here how R 1ρ MAS relaxation dispersion NMR can explore microsecond-to-millisecond motions. Progress in instrumentation, isotope labeling, and pulse sequence design has paved the way for quantitative analyses of even rare structural fluctuations. In addition to isotropic chemical-shift fluctuations exploited in solution-state NMR relaxation dispersion experiments, MAS NMR has a wider arsenal of observables, allowing to see motions even if the exchanging states do not differ in their chemical shifts. We demonstrate the potential of the technique for probing motions in challenging large enzymes, membrane proteins, and protein assemblies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Objectives: The addition of CT-derived fractional flow reserve (FFR-CT) increases the diagnostic accuracy of coronary CT angiography (CCTA). We assessed the impact of FFR-CT in routine clinical practice on clinical decision-making and patient prognosis in patients suspected of stable coronary artery disease (CAD)., Methods: This retrospective, single-center study compared a cohort that received CCTA with FFR-CT to a historical cohort that received CCTA before FFR-CT was available. We assessed the clinical management decisions after FFR-CT and CCTA and the rate of major adverse cardiac events (MACEs) during the 1-year follow-up using chi-square tests for independence. Kaplan-Meier curves were used to visualize the occurrence of safety outcomes over time., Results: A total of 360 patients at low to intermediate risk of CAD were included, 224 in the CCTA only group, and 136 in the FFR-CT group. During follow-up, 13 MACE occurred in 12 patients, 9 (4.0%) in the CCTA group, and three (2.2%) in the FFR-CT group. Clinical management decisions differed significantly between both groups. After CCTA, 60 patients (26.5%) received optimal medical therapy (OMT) only, 115 (51.3%) invasive coronary angiography (ICA), and 49 (21.9%) single positron emission CT (SPECT). After FFR-CT, 106 patients (77.9%) received OMT only, 27 (19.9%) ICA, and three (2.2%) SPECT (p < 0.001 for all three options). The revascularization rate after ICA was similar between groups (p = 0.15). However, patients in the CCTA group more often underwent revascularization (p = 0.007)., Conclusion: Addition of FFR-CT to CCTA led to a reduction in (invasive) diagnostic testing and less revascularizations without observed difference in outcomes after 1 year., Key Points: • Previous studies have shown that computed tomography-derived fractional flow reserve improves the accuracy of coronary computed tomography angiography without changes in acquisition protocols. • This study shows that use of computed tomography-derived fractional flow reserve as gatekeeper to invasive coronary angiography in patients suspected of stable coronary artery disease leads to less invasive testing and revascularization without observed difference in outcomes after 1 year. • This could lead to a significant reduction in costs, complications and (retrospectively unnecessary) usage of diagnostic testing capacity, and a significant increase in patient satisfaction., (© 2023. The Author(s).)

Aromatic side chains are important reporters of the plasticity of proteins, and often form important contacts in protein-protein interactions. We studied aromatic residues in the two structurally homologous cross-β amyloid fibrils HET-s, and HELLF by employing a specific isotope-labeling approach and magic-angle-spinning NMR. The dynamic behavior of the aromatic residues Phe and Tyr indicates that the hydrophobic amyloid core is rigid, without any sign of "breathing motions" over hundreds of milliseconds at least. Aromatic residues exposed at the fibril surface have a rigid ring axis but undergo ring flips on a variety of time scales from nanoseconds to microseconds. Our approach provides direct insight into hydrophobic-core motions, enabling a better evaluation of the conformational heterogeneity generated from an NMR structural ensemble of such amyloid cross-β architecture., (© 2023 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Background: In patients with stable coronary artery disease (CAD), revascularisation decisions are based mainly on the visual grading of the severity of coronary stenosis on invasive coronary angiography (ICA). However, invasive fractional flow reserve (FFR) is the current standard to determine the haemodynamic significance of coronary stenosis. Non-invasive and less-invasive imaging techniques such as computed-tomography-derived FFR (FFR-CT) and angiography-derived FFR (QFR) combine both anatomical and functional information in complex algorithms to calculate FFR., Trial Design: The iCORONARY trial is a prospective, multicentre, non-inferiority randomised controlled trial (RCT) with a blinded endpoint evaluation. It investigates the costs, effects and outcomes of different diagnostic strategies to evaluate the presence of CAD and the need for revascularisation in patients with stable angina pectoris who undergo coronary computed tomography angiography. Those with a Coronary Artery Disease-Reporting and Data System (CAD-RADS) score between 0-2 and 5 will be included in a prospective registry, whereas patients with CAD-RADS 3 or 4A will be enrolled in the RCT. The RCT consists of three randomised groups: (1) FFR-CT-guided strategy, (2) QFR-guided strategy or (3) standard of care including ICA and invasive pressure measurements for all intermediate stenoses. The primary endpoint will be the occurrence of major adverse cardiac events (death, myocardial infarction and repeat revascularisation) at 1 year., Clinicaltrials: gov-identifier: NCT04939207., Conclusion: The iCORONARY trial will assess whether a strategy of FFR-CT or QFR is non-inferior to invasive angiography to guide the need for revascularisation in patients with stable CAD. Non-inferiority to the standard of care implies that these techniques are attractive, less-invasive alternatives to current diagnostic pathways., (© 2023. The Author(s).)

Aims: Severe acute respiratory syndrome coronavirus-2 infection causes COVID-19, which in severe cases evokes life-threatening acute respiratory distress syndrome (ARDS). Transcriptome signatures and the functional relevance of non-vascular cell types (e.g. immune and epithelial cells) in COVID-19 are becoming increasingly evident. However, despite its known contribution to vascular inflammation, recruitment/invasion of immune cells, vascular leakage, and perturbed haemostasis in the lungs of severe COVID-19 patients, an in-depth interrogation of the endothelial cell (EC) compartment in lethal COVID-19 is lacking. Moreover, progressive fibrotic lung disease represents one of the complications of COVID-19 pneumonia and ARDS. Analogous features between idiopathic pulmonary fibrosis (IPF) and COVID-19 suggest partial similarities in their pathophysiology, yet, a head-to-head comparison of pulmonary cell transcriptomes between both conditions has not been implemented to date., Methods and Results: We performed single-nucleus RNA-sequencing on frozen lungs from 7 deceased COVID-19 patients, 6 IPF explant lungs, and 12 controls. The vascular fraction, comprising 38 794 nuclei, could be subclustered into 14 distinct EC subtypes. Non-vascular cell types, comprising 137 746 nuclei, were subclustered and used for EC-interactome analyses. Pulmonary ECs of deceased COVID-19 patients showed an enrichment of genes involved in cellular stress, as well as signatures suggestive of dampened immunomodulation and impaired vessel wall integrity. In addition, increased abundance of a population of systemic capillary and venous ECs was identified in COVID-19 and IPF. COVID-19 systemic ECs closely resembled their IPF counterparts, and a set of 30 genes was found congruently enriched in systemic ECs across studies. Receptor-ligand interaction analysis of ECs with non-vascular cell types in the pulmonary micro-environment revealed numerous previously unknown interactions specifically enriched/depleted in COVID-19 and/or IPF., Conclusions: This study uncovered novel insights into the abundance, expression patterns, and interactomes of EC subtypes in COVID-19 and IPF, relevant for future investigations into the progression and treatment of both lethal conditions., Competing Interests: Conflict of interest: A.D. received payments from FMC Belgium, and has a leadership/fiduciary role in the Belgian Society of Pathology (non-profit), and European Society of Pathology Nephropathology working group (non-profit). B.T. has a consulting role for ONO pharmaceutical and owns 10X genomics stocks. B.W. received payments from Hologic. L.J.C. received a research grant and consulting fees from MEDTRONIC. S.F. received support from Pfizer for congress attendance. W.W. received research grants and payment for lectures from Roche and Boehringer Ingelheim and a research grant from Galapagos. S.V. received consulting fees from Therakos and Boehringer Ingelheim. J.W. received investigator-initiated grants, consulting fees, speaker fees, and travel grants from Pfizer and Gilead., (© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Endothelial cells (ECs) constitute the inner lining of vascular beds in mammals and are crucial for homeostatic regulation of blood vessel physiology, but also play a key role in pathogenesis of many diseases, thereby representing realistic therapeutic targets. However, it has become evident that ECs are heterogeneous, encompassing several subtypes with distinct functions, which makes EC targeting and modulation in diseases challenging. The rise of the new single-cell era has led to an emergence of studies aimed at interrogating transcriptome diversity along the vascular tree, and has revolutionized our understanding of EC heterogeneity from both a physiological and pathophysiological context. Here, we discuss recent landmark studies aimed at teasing apart the heterogeneous nature of ECs. We cover driving (epi)genetic, transcriptomic, and metabolic forces underlying EC heterogeneity in health and disease, as well as current strategies used to combat disease-enriched EC phenotypes, and propose strategies to transcend largely descriptive heterogeneity towards prioritization and functional validation of therapeutically targetable drivers of EC diversity. Lastly, we provide an overview of the most recent advances and hurdles in single EC OMICs., Competing Interests: Conflict of interest: P.C. is named as an inventor on patent applications related to results discussed in this review., (© The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Probing the dynamics of aromatic side chains provides important insights into the behavior of a protein because flips of aromatic rings in a protein's hydrophobic core report on breathing motion involving a large part of the protein. Inherently invisible to crystallography, aromatic motions have been primarily studied by solution NMR. The question how packing of proteins in crystals affects ring flips has, thus, remained largely unexplored. Here we apply magic-angle spinning NMR, advanced phenylalanine 1 H- 13 C/ 2 H isotope labeling and MD simulation to a protein in three different crystal packing environments to shed light onto possible impact of packing on ring flips. The flips of the two Phe residues in ubiquitin, both surface exposed, appear remarkably conserved in the different crystal forms, even though the intermolecular packing is quite different: Phe4 flips on a ca. 10-20 ns time scale, and Phe45 are broadened in all crystals, presumably due to µs motion. Our findings suggest that intramolecular influences are more important for ring flips than intermolecular (packing) effects., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Author(s).)

Tumor vessel co-option (VCO) is a non-angiogenic vascularization mechanism that is a possible cause of resistance to anti-angiogenic therapy (AAT). Multiple tumors are hypothesized to primarily rely on growth factor signaling-induced sprouting angiogenesis, which is often inhibited during AAT. During VCO however, tumors invade healthy tissues by hijacking pre-existing blood vessels of the host organ to secure their blood and nutrient supply. Although VCO has been described in the context of AAT resistance, the molecular mechanisms underlying this process and the profile and characteristics of co-opted vascular cell types (endothelial cells (ECs) and pericytes) remain poorly understood, resulting in the lack of therapeutic strategies to inhibit VCO (and to overcome AAT resistance). In the past few years, novel next-generation technologies (such as single-cell RNA sequencing) have emerged and revolutionized the way of analyzing and understanding cancer biology. While most studies utilizing single-cell RNA sequencing with focus on cancer vascularization have centered around ECs during sprouting angiogenesis, we propose that this and other novel technologies can be used in future investigations to shed light on tumor EC biology during VCO. In this review, we summarize the molecular mechanisms driving VCO known to date and introduce the models used to study this phenomenon to date. We highlight VCO studies that recently emerged using sequencing approaches and propose how these and other novel state-of-the-art methods can be used in the future to further explore ECs and other cell types in the VCO process and to identify potential vulnerabilities in tumors relying on VCO. A better understanding of VCO by using novel approaches could provide new answers to the many open questions, and thus pave the way to develop new strategies to control and target tumor vascularization., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Cuypers, Truong, Becker, Saavedra-García and Carmeliet.)

Antibiotics that use novel mechanisms are needed to combat antimicrobial resistance 1-3 . Teixobactin 4 represents a new class of antibiotics with a unique chemical scaffold and lack of detectable resistance. Teixobactin targets lipid II, a precursor of peptidoglycan 5 . Here we unravel the mechanism of teixobactin at the atomic level using a combination of solid-state NMR, microscopy, in vivo assays and molecular dynamics simulations. The unique enduracididine C-terminal headgroup of teixobactin specifically binds to the pyrophosphate-sugar moiety of lipid II, whereas the N terminus coordinates the pyrophosphate of another lipid II molecule. This configuration favours the formation of a β-sheet of teixobactins bound to the target, creating a supramolecular fibrillar structure. Specific binding to the conserved pyrophosphate-sugar moiety accounts for the lack of resistance to teixobactin 4 . The supramolecular structure compromises membrane integrity. Atomic force microscopy and molecular dynamics simulations show that the supramolecular structure displaces phospholipids, thinning the membrane. The long hydrophobic tails of lipid II concentrated within the supramolecular structure apparently contribute to membrane disruption. Teixobactin hijacks lipid II to help destroy the membrane. Known membrane-acting antibiotics also damage human cells, producing undesirable side effects. Teixobactin damages only membranes that contain lipid II, which is absent in eukaryotes, elegantly resolving the toxicity problem. The two-pronged action against cell wall synthesis and cytoplasmic membrane produces a highly effective compound targeting the bacterial cell envelope. Structural knowledge of the mechanism of teixobactin will enable the rational design of improved drug candidates., (© 2022. The Author(s).)

Background: The work-up for transcatheter aortic valve replacement (TAVR) currently uses computed tomography to evaluate the annulus diameter and peripheral vascular access plus invasive coronary angiography (ICA) to assess significant coronary artery disease (CAD). ICA might partially be redundant with the use of coronary computed tomography angiography (CCTA). Prior studies found an improvement of the diagnostic accuracy of CCTA with the use of computed tomography-derived fractional flow reserve (CT-FFR)., Objectives: The aim of this study was to assess the diagnostic performance of CT-FFR for the diagnosis of CAD in the work-up for TAVR., Methods: Consecutive patients with severe symptomatic aortic valve stenosis who underwent TAVR work-up between 2015 and 2019 were included in this retrospective cross-sectional study. All patients underwent CCTA and ICA within 3 months, and the diagnostic performance of both CCTA and CT-FFR was assessed using ICA as the reference., Results: Seventy-six of the 338 patients included in the analysis had ≥1 significant coronary stenosis on ICA. The sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy per patient were 76.9%, 64.5%, 34.0%, 92.1%, and 66.9% for CCTA and 84.6%, 88.3%, 63.2%, 96.0%, and 87.6% for CT-FFR. The area under the receiver-operating characteristic curve was significantly different between CCTA and CT-FFR (0.84 vs 0.90, P = 0.02). A CT-FFR-guided approach could avoid ICA in 57.1% versus 43.6% of patients using CCTA., Conclusions: CT-FFR significantly improves the diagnostic accuracy of CCTA without additional testing and increases the proportion of patients in whom ICA could have been safely avoided. It has the potential to be integrated in the current clinical work-up for TAVR for diagnosing stable CAD requiring treatment., Competing Interests: Funding Support and Author Disclosures The authors have reported that they have no relationships relevant to the contents of this paper to disclose., (Copyright © 2022 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.)

Control of posttranscriptional mRNA decay is a crucial determinant of cell homeostasis and differentiation. mRNA lifetime is governed by cis-regulatory elements in their 3' untranslated regions (UTR). Despite ongoing progress in the identification of cis elements we have little knowledge about the functional and structural integration of multiple elements in 3'UTR regulatory hubs and their recognition by mRNA-binding proteins (RBPs). Structural analyses are complicated by inconsistent mapping and prediction of RNA fold, by dynamics, and size. We here, for the first time, provide the secondary structure of a complete mRNA 3'UTR. We use NMR spectroscopy in a divide-and-conquer strategy complemented with SAXS, In-line probing and SHAPE-seq applied to the 3'UTR of Ox40 mRNA, which encodes a T-cell co-receptor repressed by the protein Roquin. We provide contributions of RNA elements to Roquin-binding. The protein uses its extended bi-modal ROQ domain to sequentially engage in a 2:1 stoichiometry with a 3'UTR core motif. We observe differential binding of Roquin to decay elements depending on their structural embedment. Our data underpins the importance of studying RNA regulation in a full sequence and structural context. This study serves as a paradigm for an approach in analysing structured RNA-regulatory hubs and their binding by RBPs., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Skin wound repair is essential for organismal survival and failure of which leads to non-healing wounds, a leading health issue worldwide. However, mechanistic understanding of chronic wounds remains a major challenge due to lack of appropriate genetic mouse models. αSMA + myofibroblasts, a unique class of dermal fibroblasts, are associated with cutaneous wound healing but their precise function remains unknown. We demonstrate that genetic depletion of αSMA + myofibroblasts leads to pleiotropic wound healing defects, including lack of reepithelialization and granulation, dampened angiogenesis, and heightened hypoxia, hallmarks of chronic non-healing wounds. Other wound-associated FAP + and FSP1 + fibroblasts do not exhibit such dominant functions. While type I collagen (COL1) expressing cells play a role in the repair process, COL1 produced by αSMA + myofibroblasts is surprisingly dispensable for wound repair. In contrast, we show that β1 integrin from αSMA + myofibroblasts, but not TGFβRII, is essential for wound healing, facilitating contractility, reepithelization, and vascularization. Collectively, our study provides evidence for the functions of myofibroblasts in β1 integrin-mediated wound repair with potential implications for treating chronic non-healing wounds., (© 2022 The Authors.)

The vascular endothelium is characterized by a remarkable level of plasticity, which is the driving force not only of physiological repair/remodeling of adult tissues but also of pathological angiogenesis. The resulting heterogeneity of endothelial cells (ECs) makes targeting the endothelium challenging, no less because many EC phenotypes are yet to be identified and functionally inventorized. Efforts to map the vasculature at the single-cell level have been instrumental to capture the diversity of EC types and states at a remarkable depth in both normal and pathological states. Here, we discuss new EC subtypes and functions emerging from recent single-cell studies in health and disease. Interestingly, such studies revealed distinct metabolic gene signatures in different EC phenotypes, which deserve further consideration for therapy. We highlight how this metabolic targeting strategy could potentially be used to promote (for tissue repair) or block (in tumor) angiogenesis in a tissue or even vascular bed-specific manner.

Evaluation of potential immunity against the novel severe acute respiratory syndrome (SARS) coronavirus that emerged in 2019 (SARS-CoV-2) is essential for health, as well as social and economic recovery. Generation of antibody response to SARS-CoV-2 (seroconversion) may inform on acquired immunity from prior exposure, and antibodies against the SARS-CoV-2 spike protein receptor binding domain (S-RBD) are speculated to neutralize virus infection. Some serology assays rely solely on SARS-CoV-2 nucleocapsid protein (N-protein) as the antibody detection antigen; however, whether such immune responses correlate with S-RBD response and COVID-19 immunity remains unknown. Here, we generated a quantitative serological ELISA using recombinant S-RBD and N-protein for the detection of circulating antibodies in 138 serial serum samples from 30 reverse transcription PCR-confirmed, SARS-CoV-2-hospitalized patients, as well as 464 healthy and non-COVID-19 serum samples that were collected between June 2017 and June 2020. Quantitative detection of IgG antibodies against the 2 different viral proteins showed a moderate correlation. Antibodies against N-protein were detected at a rate of 3.6% in healthy and non-COVID-19 sera collected during the pandemic in 2020, whereas 1.9% of these sera were positive for S-RBD. Approximately 86% of individuals positive for S-RBD-binding antibodies exhibited neutralizing capacity, but only 74% of N-protein-positive individuals exhibited neutralizing capacity. Collectively, our studies show that detection of N-protein-binding antibodies does not always correlate with presence of S-RBD-neutralizing antibodies and caution against the extensive use of N-protein-based serology testing for determination of potential COVID-19 immunity.

The mechanistic contributions of cancer-associated fibroblasts (CAFs) in breast cancer progression remain to be fully understood. While altered glucose metabolism in CAFs could fuel cancer cells, how such metabolic reprogramming emerges and is sustained needs further investigation. Studying fibroblasts isolated from patients with benign breast tissues and breast cancer, in conjunction with multiple animal models, we demonstrate that CAFs exhibit a metabolic shift toward lactate and pyruvate production and fuel biosynthetic pathways of cancer cells. The depletion or suppression of the lactate production of CAFs alter the tumor metabolic profile and impede tumor growth. The glycolytic phenotype of the CAFs is in part sustained through epigenetic reprogramming of HIF-1α and glycolytic enzymes. Hypoxia induces epigenetic reprogramming of normal fibroblasts, resulting in a pro-glycolytic, CAF-like transcriptome. Our findings suggest that the glucose metabolism of CAFs evolves during tumor progression, and their breast cancer-promoting phenotype is partly mediated by oxygen-dependent epigenetic modifications., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

Purpose: Serial longitudinal enumeration of circulating tumor cells (CTCs) has shown its prognostic value on progression-free survival and overall survival (OS) in patients with stage IV breast cancer. This study prospectively evaluated the role of CTCs as a prognostic marker during further progression of metastatic breast cancer (MBC)., Methods: Among 476 MBC patients recruited between 2010 and 2015, the 103 patients with a known CTC status at baseline (CTC BL ) and within 4 weeks of tumor progression (CTC PD ) were included. Progressive disease (PD) was defined according to the Response Evaluation Criteria in Solid Tumors (RECIST, version 1.1). Using the CellSearch method, < 5 and ≥ 5 CTCs per 7.5 ml blood were determined as negative and positive, respectively. A shift in CTC status from baseline to progression ([Formula: see text] to [Formula: see text] and vice versa) was considered as alternating Kinetics BL-PD ., Results: Median follow-up was 29.9 [21.2, 40.0] months. CTC PD positivity (37%, n = 38) was associated with a significantly shorter OS than CTC PD negativity (8.0 [5.1, 10.9] vs 22.6 [15.3, 39.8] months; P < 0.001). Alternating Kinetics BL-PD was observed in 24% of the patients. This significantly changed the OS prediction of [Formula: see text] patients ([Formula: see text] vs [Formula: see text], 11.4 [9.7, not available (NA)] vs. 7.6 [4.4, 11.5] months; P = 0.044) and [Formula: see text] patients ([Formula: see text] vs. [Formula: see text], 8.4 [4.0, NA] vs. 22.6 [18.9, NA] months, respectively; P < 0.001). Prediction of survival was significantly improved (P = 0.002) by adding CTC PD status to clinicopathological characteristics and CTC BL status., Conclusions: CTC status upon further disease progression is a prognostic factor that could significantly improve well-established models. Thus, it represents a potential additional instrument supporting treatment decision.

On endothelial cells, the TGFβ/BMP-9 coreceptor, endoglin, emerged as a promising antiangiogenic target in colorectal cancer. Its promiscuous expression by cancer-associated fibroblasts (CAF) emerges to facilitate metastasis. The multitarget impact of neutralizing endoglin may offer added benefit in controlling colorectal cancer. See related article by Paauwe et al., p. 6331 ., (©2018 American Association for Cancer Research.)

Deregulated transforming growth factor-β (TGFβ) signaling is a common feature of many epithelial cancers. Deletion of TGFβ receptor type 2 (TGFBR2) in fibroblast specific protein-1 (FSP1)-positive stromal cells induces squamous cell carcinoma in the murine forestomach, implicating fibroblast-derived hepatocyte growth factor (HGF) as the major driver of the epithelium carcinogenesis. Prior to cancer development, hyperproliferative FSP1 + fibroblasts lacking TGFBR2 accumulate in the forestomach, disrupting the regulatory signaling cross-talk with the forestomach epithelium. Here, concurrent loss in TGFBR2 and SMAD4 completely abrogates the development of forestomach cancer. Bone morphogenic protein-7 (BMP7) was highly upregulated in forestomach cancer tissue, activating Smad1/5/8 signaling, cell proliferation, and HGF production in TGFBR2 -deficient FSP1 + fibroblasts. This stimulation by BMP7 was lost in the combined TGFBR2 and SMAD4 double knockout fibroblasts, which included a profound decrease in HGF expression. Thus, Smad4-mediated signaling is required to initiate epithelial carcinogenesis subsequent to TGFBR2 deletion in FSP1 + fibroblasts. Implications: These findings reveal a complex cross-talk between epithelial cells and the stroma, wherein Smad4 is required to elicit squamous cell carcinomas in the forestomach of mice with TGFBR2 -deficient stromal cells. Mol Cancer Res; 16(10); 1568-78. ©2018 AACR ., (©2018 American Association for Cancer Research.)

Over the past two years, the business community of Monroe County, which includes Rochester, New York, has been engaging in a collaborative to improve outcomes for people with high blood pressure. As the employers examined the costs of care in the community, they recognized two important factors. First, the costs of care for the uninsured, the underinsured, and the Medicare population influence the business community's cost of care. Second, trying to redesign care just for their employees alone was not effective. This project is unique in that the stimulus and funding for community-wide action comes from the business community. They saw beyond the often unsuccessful short-term cost reduction programs and joined with a community-focused organization, the Finger Lakes Health Systems Agency, to construct a multi-year, multi-faceted intervention designed to encourage practice redesign and an invigorated community commitment to partnership and accountability. This report describes the process to date and hopefully will stimulate conversations about mechanisms to encourage similar collaboration within other communities., (© 2012 Wiley Periodicals, Inc.)

Aberrant protein processing with tissue deposition is associated with many common neurodegenerative disorders; however, the complex interplay of genetic and environmental factors has made it difficult to decipher the sequence of events linking protein aggregation with clinical disease. Substantial progress has been made toward understanding the pathophysiology of prototypical conformational diseases and protein polymerization in the superfamily of serine proteinase inhibitors (serpins). Here we describe a new disease, familial encephalopathy with neuroserpin inclusion bodies, characterized clinically as an autosomal dominantly inherited dementia, histologically by unique neuronal inclusion bodies and biochemically by polymers of the neuron-specific serpin, neuroserpin. We report the cosegregation of point mutations in the neuroserpin gene (PI12) with the disease in two families. The significance of one mutation, S49P, is evident from its homology to a previously described serpin mutations, whereas that of the other, S52R, is predicted by modelling of the serpin template. Our findings provide a molecular mechanism for a familial dementia and imply that inhibitors of protein polymerization may be effective therapies for this disorder and perhaps for other more common neurodegenerative diseases.